US 2008O148432A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2008/0148432 A1 Abad (43) Pub. Date: Jun. 19, 2008

(54) TRANSGENIC PLANTS WITH ENHANCED Publication Classification AGRONOMIC TRAITS (51) Int. Cl. AOIH 5/00 (2006.01) CI2N 5/82 (2006.01) (76) Inventor: Mark Scott Abad, Webster Grove, CI2N 5/04 (2006.01) MO (US) (52) U.S. Cl...... 800/279: 800/281; 435/419,435/468; 8OO/320.1 Correspondence Address: (57)57 ABSTRACT MONSANTO COMPANY This invention provides transgenic plant cells with recombi 800 N. LINDBERGHBLVD, ATTENTION: GAIL nant DNA for expression of proteins that are useful for P. WUELLNER, IP PARALEGAL (E2NA) imparting enhanced agronomic trait(s) to transgenic crop ST. LOUIS MO 631.67 9 plants. This invention also provides transgenic plants and 9 progeny seed comprising the transgenic plant cells where the plants are selected for having an enhanced trait selected from the group of traits consisting of enhanced water use effi (21) Appl. No.: 11/374,300 ciency, enhanced cold tolerance, increased yield, enhanced nitrogen use efficiency, enhanced seed protein and enhanced seed oil. Also disclosed are methods for manufacturing trans (22) Filed: Dec. 21, 2005 genic seed and plants with enhanced traits. Patent Application Publication Jun. 19, 2008 Sheet 1 of 3 US 2008/0148432 A1

Figure 1. 41905 ------127 O2 ------721 O ------24286 MTGFGLLVOTISSGASGHIDEGAADEEEARWSSWRRKELRGGAVVAHGTGWCACGTTSTP 6881 ------MHTSVICPLPvPRMAVFSASKAVSDESAFRVAGRVFVFVSSRL 5 O247 ------9016 ------205 ------81.07 ------45.508 ------28787 ------17886 ------28339 ------consensus ------

MVKKKPAG MVKKKPAG MVKKKEAG IWRSRSGNTTPAPLWRNELERLGLTTRMDTDIGIID-FGFSLLPFITILLIDMESWTLSL VPNTRTRHADTRIVPPNFCSEISSSRLISLFOSITSSNLYPKPYTGGRGMVKKAVSTAPA

------as a ------MVKKAVSTAPA MVAKKPAAAAAA

------MVRASSSKKGGSKGGDKD

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------r m r ------XXX XXXXXXXXXXXX XXX

DAEADERRRLRSLAFSNGLLQRGEPAAPRSALAPS-TAVSRLOGRDIVRRGGQRKSRFLF DAEADERRRLRSLAFSNGLLQRGEPAAPRSALAPS-TAVSRLOGRDIVRRGGORKSRFLF DAEADERRRLRSLAFSNGLLORGEPAAPRSALAPS-TAVSRLQGRDIVRRGGQRKSRFLF PPPFPPRSSASSLTSACQRREEIELSCKRHVNAMQGRAQRVVOGRDIVRRGGQRKSRFLF DAEADERRRL, RSLAFSNGLLQRGDPAAPRA PLAPA-AAVTRLQGRDVVRRGGQRKSRYLF DAEADERRRLRSLAFSNGL LORGDPAAPRA PLAPA-AAVTRLQGRDVVRRGGQRKSRILF GPEAEERRER LRSLAFSKGL LORGEP&APRAALPPS-GAVARLQGRDIVRPRGORRGRFLF DAESKQRKRLKTLALDNQLLSD-SPAKSHSSLKPS-KQVLKHHGT DIIRKSORK-NRFLF DAESKQRix RLKTLALDNQLLSD-SPAKSHSSLKPT-KQVLKHHGT DIIRKSQRK-NRFLF DAESKQRKRLKTLA LDNQLLSD-SPAKSHSS LKPS - KOVLKHHGT DIIRKSORK-NRFLF OPESKQRKRLKSLACKL LSD-SPSRCLSS. KPS-KOVLKHHGCDI IRKSORK-NRFLF NPETLERKRLKSLA ISNKILSE-T PARSSVF Liy PS-SVVAKHHGKDIIKKSQRKSCR; LF NPETLERKRLKSLA ISNKILSE-TPARSSVHLNPS-SVVAKHHGKDI IKKSQRKSCRYLF XXe XXXRX riXS Lax:n:ll xxxxpaXX:XX lxpx-xxv XXXXGXDixir xxxx xxxRXLF

SFPGLLAPAAAASGGRVGELADLGTKNPLLYLDFPQGRMKLLGTHVYPKNKYLTLOX--- SFPGLLAPAAAASGGRVGELADLGTKNPLLYLDFPQGRMKLLGTHVYPKNKYLTLOX--- SFPGLLAPAAAASGGRVGELADLGTKNPLLYLDFPOGRMKLLGTHVYPKNKYLTLOMSRS SFPGLLAPAAAASGGRVGELADLGTKNPLLYLDFPOGRMKLLGTHVYPKNKYLTLOMSRS SFPGLLAPAASG--GRVGELADLGTKNPLLYLEFPQGRMKLFGTHVYPKNKYLTLQMTRS SFPGLLAPAASG--GRVGELADLGTKNPLLYLEFPQGRMKLFGTHVYPKNKYLTLQMTRS SFPGLLAPAAAG--GRVGELADLGTKNPVLYLEFPOGRMKLLGTHVYPKNKYLTLOMTRS SFPGLLAPIS AA- - -TIGDLDRLSTKNPVLYLNFPQGRMKLFGTILYPKNRYLTLOFSRG SFPGLLAPIS AA- - -TIGDLDRLSTKNPVLYLNFPOGRMKLFGTILYPKNRYLTLOFSRG

Patent Application Publication Jun. 19, 2008 Sheet 3 of 3 US 2008/O148432 A1

DIKEKKSQWKTFNPEDPPAESLSIKKPPXLVOANLSAQVXKNQKKKRDPHSRKSPRGSPA DGNGITASASKL PEEL PAKREKLKSKDSKLVOATLSNLFKKAEEKTAGTSKAKSSSKA-- DGNGITASASKL PEEL PAKREKLKSKDSKLVOATLSNLFKKAEEKTAGTSKAKSSSKA-- DGNGVTASASKLPTELPAKKEKPKSKDSKLVOATLSNLFKKAEEKTAGTSKAKSSSKA-- QADITTASTGKLPTELPAKKEKSNSKDGKLVOATWANLFKKAEQKTVGTSKAKSXXKAAO TVVFDLDDEDDAPVDQPAKINTES ASRSKSKEWSQSASASASTEVKSSNRGSLVOATIST TVVFDLDDEDDAPVDQPAKKNNES ASRSKCKGRCLNLLQRVHLLKLSPVIVVHLFRLLYP dXXXXXXSXXXXXXXXXXXXXXXX sixxxkxxxxxxxxxxxxxxxxxxxxxxxkxxxxxxx

EKHPTGKKSAGRSQKRRKTQVEDDKIEVLSSSSQVVPSSQLILTAYDRITTWTMIAMRTG EKHPTGKKSAGRSQKKRKTQVEDDEIEVLSSSSQDNNVDDDSDEDWAE------EKHPTGKKSGKCSSKSVVRSI ------EKHPTGKKSGKCSSKSVVRSV------AKOPAGIKKVSGTRGKKKPKVGEDEIEELSSSSODNDADDDSDEDWAE------

YKGPPCSKAGGTSPPKTSHP------

KK------IFKKWEEKTTPRSSRKSPSSKAYGOKSOPAGSKRKIDLDEGSKKRARKTTDKDPGKKIKA HYSRKWKKRKLQEWRGNLHLONLLARSHSLLVOKERLIWLNFLFLFFIS ------XXXXXXX XXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXX ------

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TRANSGENC PLANTS WITHENHANCED from a population are required to identify those transgenic AGRONOMIC TRAITS events that are characterized by the enhanced agronomic trait.

CROSS REFERENCE TO RELATED SUMMARY OF THE INVENTION APPLICATIONS 0007. This invention employs recombinant DNA for expression of proteins that are useful for imparting enhanced 0001. This application claims benefit under 35USC S 119 agronomic traits to the transgenic plants. Recombinant DNA (e) of U.S. provisional application Ser. No. 60/638,099, filed in this invention is provided in a construct comprising a Dec. 21, 2004, herein incorporated by reference. promoter that is functional in plant cells and that is operably linked to DNA that encodes a protein having at least one INCORPORATION OF SEQUENCE LISTING amino acid domain in a sequence that exceeds the Pfam gathering cutoff for amino acid sequence alignment with a 0002. Two copies of the sequence listing (Copy 1 and protein domain family identified by a Pfam name in the group Copy 2) and a computer readable form (CRF) of the sequence of Pfam domain names as identified in Table 12. In more listing, all on CD-Rs, each containing the text file named specific embodiments of the invention the protein expressed 38-21 (53720)D seqListing..txt, which is 192,434,176 bytes in plant cells has an amino acid sequence with at least 90% (measured in MS-WINDOWS) and was created on Dec. 21, identity to a consensus amino acid sequence in the group of 2005, are herein incorporated by reference. consensus amino acid sequences consisting of the consensus amino acid sequence constructed for SEQID NO: 742 and INCORPORATION OF COMPUTER PROGRAM homologs thereof listed in Table 2 through the consensus LISTING amino acid sequence constructed for SEQID NO: 1482 and homologs thereof listed in Table 2. In even more specific 0003 Computer Program Listing folders hmmer-2.3.2 embodiments of the invention the protein expressed in plant and 347pfamDir are contained on a compact disc and is cells is a protein selected from the group of proteins identified hereby incorporated herein by reference in their entirety. in Table 1. Folder hmmer-2.3.2 contains the source code and other asso 0008. Other aspects of the invention are specifically ciated file for implementing the HMMer software for Pfam directed to transgenic plant cells comprising the recombinant analysis. Folder 347pfamIDir contains 347 Pfam Hidden DNA of the invention, transgenic plants comprising a plural Markov Models. Both folders were created on the disk on ity of Such plant cells, progeny transgenic Seed, embryo and Dec. 21, 2005, having a total size of 53,825,536 bytes (mea transgenic pollen from Such plants. Such plant cells are sured in MS-WINDOWS). selected from a population of transgenic plants regenerated from plant cells transformed with recombinant DNA and that express the protein by Screening transgenic plants in the INCORPORATION OF TABLE population for an enhanced trait as compared to control plants that do not have said recombinant DNA, where the enhanced 0004 Two copies of Tables 2 on CD-Rs, each containing trait is selected from group of enhanced traits consisting of the file named 38-21 (53720)D table2.txt, which is 24,334, enhanced water use efficiency, enhanced cold tolerance, 336 bytes (measured in MS-WINDOWS), was created on increased yield, enhanced nitrogen use efficiency, enhanced Dec. 21, 2005, and comprises 132 pages when viewed in MS seed protein and enhanced seed oil. Word, are herein incorporated by reference. 0009. In yet another aspect of the invention the plant cells, plants, seeds, embryo and pollen further comprise DNA FIELD OF THE INVENTION expressing a protein that provides tolerance from exposure to an herbicide applied at levels that are lethal to a wild type of 0005 Disclosed herein are inventions in the field of plant said plant cell. Such tolerance is especially useful not only as genetics and developmental biology. More specifically, the a advantageous trait in Such plants but is also useful in a present inventions provide plant cells with recombinant DNA selection step in the methods of the invention. In aspects of for providing an enhanced trait in a transgenic plant, plants the invention the agent of Such herbicide is a glyphosate, comprising Such cells, seed and pollen derived from Such dicamba, or glufosinate compound. plants, methods of making and using such cells, plants, seeds 0010 Yet other aspects of the invention provide transgenic and pollen. plants which are homozygous for the recombinant DNA and transgenic seed of the invention from corn, soybean, cotton, BACKGROUND OF THE INVENTION canola, alfalfa, wheat or rice plants. In other important embodiments for practice of various aspects of the invention 0006 Transgenic plants with improved agronomic traits in Argentina the recombinant DNA is provided in plant cells Such as yield, environmental stress tolerance, pest resistance, derived from corn lines that that are and maintain resistance to herbicide tolerance, improved seed compositions, and the like the Mal de Rio Cuarto virus or the Puccina sorghi fungus or are desired by both farmers and consumers. Although consid both. erable efforts in plant breeding have provided significant 0011. This invention also provides methods for manufac gains in desired traits, the ability to introduce specific DNA turing non-natural, transgenic seed that can be used to pro into plant genomes provides further opportunities for genera duce a crop of transgenic plants with an enhanced trait result tion of plants with improved and/or unique traits. Merely ing from expression of stably-integrated, recombinant DNA introducing recombinant DNA into a plant genome doesn't for expressing a protein having at least one domain of amino always produce a transgenic plant with an enhanced agro acids in a sequence that exceeds the Pfamgathering cutoff for nomic trait. Methods to select individual transgenic events amino acid sequence alignment with a protein domain family US 2008/0148432 A1 Jun. 19, 2008

identified by a Pfam name in the group of Pfam names iden invention also provides methods of growing a corn, cotton or tified in Table 12. More specifically the method comprises (a) Soybean crop without added nitrogen fertilizer comprising screening a population of plants for an enhanced trait and a planting seed having plant cells of the invention which are recombinant DNA, where individual plants in the population selected for enhanced nitrogen use efficiency. can exhibit the trait at a level less than, essentially the same as or greater than the level that the trait is exhibited in control DETAILED DESCRIPTION OF THE INVENTION plants which do not express the recombinant DNA, (b) select 0015. As used herein a “plant cell means a plant cell that ing from the population one or more plants that exhibit the is transformed with stably-integrated, non-natural, recombi trait at a level greater than the level that said trait is exhibited nant DNA, e.g. by Agrobacterium-mediated transformation in control plants, (c) verifying that the recombinant DNA is or by baombardment using microparticles coated with recom stably integrated in said selected plants, (d) analyzing tissue binant DNA or other means. A plant cell of this invention can of a selected plant to determine the production of a protein be an originally-transformed plant cell that exists as a micro having the function of a protein encoded by nucleotides in a organism or as a progeny plant cell that is regenerated into sequence of one of SEQID NO: 1-741; and (e) collecting seed differentiated tissue, e.g. into a transgenic plant with stably from a selected plant. In one aspect of the invention the plants integrated, non-natural recombinant DNA, or seed or pollen in the population further comprise DNA expressing a protein derived from a progeny transgenic plant. that provides tolerance to exposure to an herbicide applied at 0016. As used herein a “transgenic plant’ means a plant levels that are lethal to wild type plant cells and the selecting whose genome has been altered by the stable integration of is effected by treating the population with the herbicide, e.g. recombinant DNA. A transgenic plant includes a plant regen a glyphosate, dicamba, or glufosinate compound. In another erated from an originally-transformed plant cell and progeny aspect of the invention the plants are selected by identifying transgenic plants from later generations or crosses of a trans plants with the enhanced trait. The methods are especially formed plant. useful for manufacturing corn, soybean, cotton, alfalfa, wheat 0017. As used herein “recombinant DNA means DNA or rice seed. which has been a genetically engineered and constructed 0012 Another aspect of the invention provides a method outside of a cell including DNA containing naturally occur of producing hybrid corn seed comprising acquiring hybrid ring DNA or cDNA or synthetic DNA. corn seed from a herbicide tolerant corn plant which also has 0018. As used herein “consensus sequence” means an arti stably-integrated, recombinant DNA comprising a promoter ficial sequence of amino acids in a conserved region of an that is (a) functional in plant cells and (b) is operably linked to alignment of amino acid sequences of homologous proteins, DNA that encodes a protein having at least one domain of e.g. as determined by a CLUSTALW alignment of amino acid amino acids in a sequence that exceeds the Pfam gathering sequence of homolog proteins. cutoff for amino acid sequence alignment with a protein 0019. As used herein “homolog” means a protein in a domain family identified by a Pfam name in the group of group of proteins that perform the same biological function, Pfam names identified in Table 12. The methods further com e.g. proteins that belong to the same Pfam and prise producing corn plants from said hybrid corn seed, that provide a common enhanced trait in transgenic plants of wherein a fraction of the plants produced from said hybrid this invention. Homologs are expressed by homologous corn seed is homozygous for said recombinant DNA, a frac genes. Homologous genes include naturally occurring alleles tion of the plants produced from said hybrid corn seed is and artificially-created variants. Degeneracy of the genetic hemizygous for said recombinant DNA, and a fraction of the code provides the possibility to substitute at least one base of plants produced from said hybrid corn seed has none of said the protein encoding sequence of a gene with a different base recombinant DNA, selecting corn plants which are homozy without causing the amino acid sequence of the polypeptide gous and hemizygous for said recombinant DNA by treating produced from the gene to be changed. Hence, a polynucle with an herbicide; collecting seed from herbicide-treated otide useful in the present invention may have any base Surviving corn plants and planting said seed to produce fur sequence that has been changed from SEQID NO: 1 through ther progeny corn plants; repeating the selecting and collect SEQID NO: 741 substitution in accordance with degeneracy ing steps at least once to produce an inbred corn line; and of the genetic code. Homologs are proteins that, when opti crossing the inbred corn line with a second corn line to pro mally aligned, have at least 60% identity, more preferably duce hybrid seed. about 70% or higher, more preferably at least 80% and even 0013 Another aspect of the invention provides a method more preferably at least 90% identity over the full length of a of selecting a plant comprising plant cells of the invention by protein identified as being associated with imparting an using an immunoreactive antibody to detect the presence of enhanced trait when expressed in plant cells. Homologs protein expressed by recombinant DNA in seed or plant tis include proteins with an amino acid sequence that has at least Sue. Yet another aspect of the invention provides anti-coun 90% identity to a consensus amino acid sequence of proteins terfeit milled seed having, as an indication of origin, a plant and homologs disclosed herein. cells of this invention. 0020 Homologs are be identified by comparison of amino 0014 Still other aspects of this invention relate to trans acid sequence, e.g. manually or by use of a computer-based genic plants with enhanced water use efficiency or enhanced tool using known homology-based search algorithms such as nitrogen use efficiency. For instance, this invention provides those commonly known and referred to as BLAST. FASTA, methods of growing a corn, cotton or soybean crop without and Smith-Waterman. A local sequence alignment program, irrigation water comprising planting seed having plant cells e.g. BLAST, can be used to search a database of sequences to of the invention which are selected for enhanced water use find similar sequences, and the Summary Expectation value efficiency. Alternatively methods comprise applying reduced (E-value) used to measure the sequence base similarity. As a irrigation water, e.g. providing up to 300 millimeters of protein hit with the best E-value for aparticular organism may ground water during the production of a corn crop. This not necessarily be an ortholog or the only ortholog, a recip US 2008/0148432 A1 Jun. 19, 2008

rocal query is used in the present invention to filter hit protein encoded by candidate DNA against the Hidden sequences with significant E-values for ortholog identifica Markov Model which characterizes the Pfam domain using tion. The reciprocal query entails search of the significant hits HMMER software, a current version of which is provided in against a database of amino acid sequences from the base the appended computer listing. Candidate proteins meeting organism that are similar to the sequence of the query protein. the gathering cutoff for the alignment of a particular Pfam are A hit is a likely ortholog, when the reciprocal query's best hit in the protein family and have cognate DNA that is useful in is the query protein itselfora protein encoded by a duplicated constructing recombinant DNA for the use in the plant cells of gene after speciation. A further aspect of the invention com this invention. Hidden Markov Model databases for use with prises functional homolog proteins that differ in one or more HMMER software in identifying DNA expressing protein in amino acids from those of disclosed protein as the result of a common Pfam for recombinant DNA in the plant cells of conservative amino acid Substitutions, for example Substitu this invention are also included in the appended computer tions are among: acidic (negatively charged) amino acids listing. The HMMER software and Pfam databases are ver Such as aspartic acid and glutamic acid; basic (positively sion 18.0 and were used to identify known domains in the charged) amino acids Such as arginine, histidine, and lysine; proteins corresponding to amino acid sequence of SEQ ID neutral polar amino acids such as glycine, serine, threonine, NO: 742 through SEQ ID NO: 1482. All DNA encoding cysteine, tyrosine, asparagine, and glutamine; neutral nonpo proteins that have scores higher than the gathering cutoff lar (hydrophobic) amino acids Such as alanine, leucine, iso disclosed in Table 12 by Pfam analysis disclosed herein can leucine, Valine, proline, phenylalanine, tryptophan, and be used in recombinant DNA of the plant cells of this inven methionine; amino acids having aliphatic side chains such as tion, e.g. for selecting transgenic plants having enhanced glycine, alanine, Valine, leucine, and isoleucine; amino acids agronomic traits. The relevant Pfams for use in this invention, having aliphatic-hydroxyl side chains such as serine and as more specifically disclosed below, are bZIP 1, bZIP 2, threonine; amino acids having amide-containing side chains Meth synt 1, Homeobox, Succ DH flav C. RWP-RK, Such as asparagine and glutamine; amino acids having aro Meth synt 2, CTP synth N, WD40, Sigma70 r2, matic side chains such as phenylalanine, tyrosine, and tryp Sigma70 r3, Fer4, Sigma70 ra. Sigma70 r1 2, CMAS, tophan; amino acids having basic side chains such as lysine, Sugar tr. Rubrerythrin, Pro dh, Ldh 1. C. START, HAT arginine, and histidine, amino acids having Sulfur-containing Pase c, Cpn10, Glycos transf 1, Glycos transf 2, Pki side chains such as cysteine and methionine; naturally con nase, KH 1, cobW. Ldh 1 N, DUF393, SecY, PCI, SRF servative amino acids such as valine-leucine, Valine-isoleu TF, IF4E. Lectin legA, Mat, Dehydrin, Lectin legEB, Ank, cine, phenylalanine-tyrosine, lysine-arginine, alanine-valine, 2-Hacid dh C, Tic22, Chal sti synt C, AA kinase, ELFV aspartic acid-glutamic acid, and asparagine-glutamine. A fur dehydrog N, HLH. Ribonuclease T2, HEM4, AT hook, ther aspect of the homologs encoded by DNA useful in the Peptidase A22B, tRNA-synt 2b, Suc Fer-like, Glyco transgenic plants of the invention are those proteins that differ transf 20, MFS 1, HMA, Ketoacyl-synt C. Steroid dh, from a disclosed protein as the result of deletion or insertion , Peptidase C1, Ion trans, Aa trans, peroxidase, of one or more amino acids in a native sequence. GAF, Cu-oxidase, ABC1, PMSR, B12D, Chromo, Lipase 0021. As used herein, “percent identity” means the extent GDSL, Ran BP1, DUF125, Lig chan, GAT. Tub, NPH3, to which two optimally aligned DNA or protein segments are BAH, GFO IDH MocA, DUF6, Orn DAP Arg deC, invariant throughout a window of alignment of components, F-box, 3 5 exonuc, NUDIX, Cyclin C, Trehalase Ca-bi, for example nucleotide sequence oramino acid sequence. An Acyltransferase, MtN3 slv, Zf-B box, PUA, AMPKBI, Pep “identity fraction’ for aligned segments of a test sequence and tidase M20, Transaldolase, ketoacyl-synt, Cyclin N. a reference sequence is the number of identical components HisKA, Ribosomal L7Ae, Methyltransf 11, Methyl that are shared by sequences of the two aligned segments transf 12, Hexapep, Ribosomal S2, Jacalin, ERp29, divided by the total number of sequence components in the MFMR, Usp, DUF641, Pyr redox dim, Auxin resp. Inhibi reference segment over a window of alignment which is the tor 129, , cnMP binding, BURP, Epimerase, smaller of the full test sequence or the full reference Ribosomal L39, Metallothio 2, Pyr redox 2, WRKY. sequence. “Percent identity” (“yo identity”) is the identity GSHPx, Kelch 1, Kelch 2, Aminotran 1, 2, ABC tran, fraction times 100. UDPGT, Cystatin, YL1, AMP-binding, NTP transferase, 0022. As used herein “Pfam” refers to a large collection of HALZ, Kunitz legume, HSP20, DUF581, FGGY N, Ami multiple sequence alignments and hidden Markov models notran 3, PHD, B56, Aminotran 5, PSI PsaE, malic, covering many common protein families, e.g. Pfam version Zf-C2H2, HEAT, UPF0057, Asn synthase, K-box, HAMP. 18.0 (August 2005) contains alignments and models for 7973 PTR2, SapB 1, Ammonium transp, SapB 2, GATase, Pyr protein families and is based on the Swissprot 47.0 and SP redox, Cu-oxidase 2, Cu-oxidase 3, Cyclotide, Asp, M20 TrEMBL 30.0 protein sequence databases. See S. R. Eddy, dimer, PA. Thiolase C, FHA, Yief N, Citrate synt, GTP “Profile Hidden Markov Models”, Bioinformatics 14:755 EFTU D2, GTP EFTU D3, PK, GATA, Thiolase N, 763, 1998. Pfam is currently maintained and updated by a Glycogen syn, WHEP-TRS, B3, EF1 GNE, FAD binding Pfam Consortium. The alignments represent some evolution 3, ComA, Remorin C, FAD binding 7, RimlD sub bind, ary conserved structure that has implications for the protein's CBS, ELFV dehydrog, YL1 C, Zf-Dof Ribosomal S11, function. Profile hidden Markov models (profile HMMs) ArfGap, GRAS, Metallophos, Annexin, Ras, NAC, Acetyl built from the Pfam alignments are useful for automatically transf 1, Ribosomal S17, NAF, DUF246, GST C, CN hy recognizing that a new protein belongs to an existing protein drolase, Na Ca ex, DUF1423, Ubie methyltran, p450, family even if the homology by alignment appears to be low. PP2C, NAM, Histone, GST N, Tubulin, 2-Hacid dh, Ribo Once one DNA is identified as encoding a protein which somal L19e, CCT, Malic M, PKC, VHS, IPK, HSF DNA imparts an enhanced trait when expressed in transgenic bind, Tubulin C, Sina, JmjC, CH, Catalase, DUF250, HMG plants, other DNA encoding proteins in the same protein box, PfkB, Yippee, DSPc, Pkinase C, UbiA, Ribosomal family are identified by querying the amino acid sequence of S27, ADH zinc N. Zip, Globin, JmjN, Cys Met Meta PP, US 2008/0148432 A1 Jun. 19, 2008

H10933 like, GH3, Bromodomain, ERO1, DAO, DUF760, cases be a progeny of a hemizygous transgenic plant line that Methyltransf 2, Gp dh C, HGTP anticodon, Methyltransf is does not contain the recombinant DNA, known as a nega 3, Aldo ket red, Thioredoxin, NimrA, SelR, LEA 5. Orn tive segregant. Arg deC. N. Polysacc synt 2, Gp dh N. Niflu N, GFO 0027. As used herein an "enhanced trait” means a charac IDH MocAC, Gamma-thionin, FBA 1, HPPase, ADH teristic of a transgenic plant that includes, but is not limited to, N, Heme oxygenase, AUX IAA, NAD binding 4. Auxin an enhance agronomic trait characterized by enhanced plant inducible, LIM, Response reg, Dirigent, E2F TDP, Di19, morphology, physiology, growth and development, yield, Alpha adaptinC2, efhand, ICL, Rieske, GTP EFTU, ARID, nutritional enhancement, disease or pest resistance, or envi adh short, Transket pyr, AA permease, TPP C. ronmental or chemical tolerance. In more specific aspects of NDK, RRM 1, Trypsin, Pro CA, Hexokinase 1, CBFD this invention enhanced trait is selected from group of NFYB HMF, Glyco hydro 38C, TPP enzyme M, TPP enhanced traits consisting of enhanced water use efficiency, enzyme N, Hexokinase 2, 3Beta HSD, DUF788, WZy C, enhanced cold tolerance, increased yield, enhanced nitrogen E1 dh, Glycolytic, RuBisCO small, ZF-HD dimer, use efficiency, enhanced seed protein and enhanced seed oil. DUF1530, PARP, Pyridoxal deC, IlvC, Ribosomal L1, In an important aspect of the invention the enhanced trait is Alpha-amylase, EB1, CorA, Sucrose synth, PGAM, IlvN. enhanced yield including increased yield under non-stress MAP1 LC3, DNA photolyase, PAD porph, Abhydrolase conditions and increased yield under environmental stress 1, Glyco hydro 16, NTF2, CobW. C. GATase 2, Cation conditions. Stress conditions may include, for example, efflux, Gln-synt C. VQ, DUF296, W2, SAM 1, SAM 2. drought, shade, fungal disease, viral disease, bacterial dis Gln-synt N. Transketolase C, PEPcase, GRIM-19, Pkinase ease, insect infestation, nematode infestation, cold tempera Tyr. DnaJ, MIP, PRA1, Trehalose PPase, Transketolase N, ture exposure, heat exposure, osmotic stress, reduced nitro LRR 2. KA1, Mpv17 PMP22, Reticulon, Trp syntA,YTH, gen nutrient availability, reduced phosphorus nutrient Aldedh, Zf-C3HC4, GIDA, Trp Tyr perm, UBA, PB1, PAS, availability and high plant density. “Yield” can be affected by Carb kinase, Zf-LSD1, CAF1, Xan ur permease. Hist many properties including without limitation, plant height, deacetyl, Cpnó0 TCP1, XET C, Ribosomal L10e, Tre pod number, pod position on the plant, number of internodes, halase, ubiquitin, Glyco hydro 38, AP2, Myb DNA-bind incidence of pod shatter, grain size, efficiency of nodulation ing, APS kinase, PBD, FAE 3-kCoA syn1, the databases and nitrogen fixation, efficiency of nutrient assimilation, for which are included in the appended computer listing. resistance to biotic and abiotic stress, carbon assimilation, 0023. As used herein “promoter” means regulatory DNA plant architecture, resistance to lodging, percent seed germi for initializing transcription. A plant promoter is a promoter nation, seedling vigor, and juvenile traits. Yield can also capable of initiating transcription in plant cells whether or not affected by efficiency of germination (including germination its origin is a plant cell, e.g. is it well known that Agrobacte in stressed conditions), growth rate (including growth rate in rium promoters are functional in plant cells. Thus, plant pro stressed conditions), ear number, seed number per ear, seed moters include promoter DNA obtained from plants, plant size, composition of seed (starch, oil, protein) and character viruses and bacteria Such as Agrobacterium and Bradyrhizo istics of seed fill. bium bacteria. Examples of promoters under developmental 0028. Increased yield of a transgenic plant of the present control include promoters that preferentially initiate tran invention can be measured in a number of ways, including test Scription in certain tissues. Such as leaves, roots, or seeds. weight, seed number per plant, seed weight, seed number per Such promoters are referred to as “tissue preferred. Promot unit area (i.e. seeds, or weight of seeds, per acre), bushels per ers that initiate transcription only in certain tissues are acre, tonnes per acre, tons per acre, kilo per hectare. For referred to as “tissue specific''. A "cell type specific pro example, maize yield may be measured as production of moter primarily drives expression in certain cell types in one shelled corn kernels per unit of production area, for example or more organs, for example, vascular cells in roots or leaves. in bushels per acre or metric tons per hectare, often reported An “inducible' or “repressible' promoter is a promoter which on a moisture adjusted basis, for example at 15.5 percent is under environmental control. Examples of environmental moisture. Increased yield may result from improved utiliza conditions that may effect transcription by inducible promot tion of key biochemical compounds, such as nitrogen, phos ers include anaerobic conditions, or certain chemicals, or the phorous and carbohydrate, or from improved responses to presence of light. Tissue specific, tissue preferred, cell type environmental stresses, such as cold, heat, drought, salt, and specific, and inducible promoters constitute the class of “non attack by pests or pathogens. Recombinant DNA used in this constitutive' promoters. A “constitutive' promoter is a pro invention can also be used to provide plants having improved moter which is active under most conditions. growth and development, and ultimately increased yield, as 0024. As used herein “operably linked' means the asso the result of modified expression of plant growth regulators or ciation of two or more DNA fragments in a DNA construct so modification of cell cycle or photosynthesis pathways. Also that the function of one, e.g. protein-encoding DNA, is con of interest is the generation of transgenic plants that demon trolled by the other, e.g. a promoter. strate enhanced yield with respect to a seed component that 0025. As used herein “expressed” means produced, e.g. a may or may not correspond to an increase in overall plant protein is expressed in a plant cell when its cognate DNA is yield. Such properties include enhancements in seed oil, seed transcribed to mRNA that is translated to the protein. molecules such as tocopherol, protein and starch, or oil par 0026. As used herein a “control plant’ means a plant that ticular oil components as may be manifest by an alterations in does not contain the recombinant DNA that expressed a pro the ratios of seed components. tein that impart an enhanced trait. A control plant is to identify 0029. A subset of the nucleic molecules of this invention and select a transgenic plant that has an enhance trait. A includes fragments of the disclosed recombinant DNA con Suitable control plant can be a non-transgenic plant of the sisting of oligonucleotides of at least 15, preferably at least 16 parental line used to generate a transgenic plant, i.e. devoid of or 17, more preferably at least 18 or 19, and even more recombinant DNA. A suitable control plant may in some preferably at least 20 or more, consecutive nucleotides. Such US 2008/0148432 A1 Jun. 19, 2008

oligonucleotides are fragments of the larger molecules having flower mosaic virus (CaMV35S), U.S. Pat. No. 5,641,876, a sequence selected from the group consisting of SEQID NO: which discloses a rice actin promoter, U.S. Patent Application 1 through SEQ ID NO: 741, and find use, for example as Publication 2002/0192813A1, which discloses 5', 3' and probes and primers for detection of the polynucleotides of the intron elements useful in the design of effective plant expres present invention. sion vectors, U.S. patent application Ser. No. 09/757,089, 0030. In some embodiments of the present invention, a which discloses a maize chloroplast aldolase promoter, U.S. dominant negative mutant of a native gene is generated to patent application Ser. No. 08/706,946, which discloses a rice achieve the desired effect. As used herein, "dominant nega glutelin promoter, U.S. patent application Ser. No. 09/757, tive mutant’ means a mutant gene whose gene 089, which discloses a maize aldolase (FDA) promoter, and adversely affects the normal, wild-type gene product within U.S. patent application Ser. No. 60/310,370, which discloses the same cell, usually by dimerizing (combining) with it. In a maize nicotianamine synthase promoter, all of which are cases of polymeric molecules, such as collagen, dominant incorporated herein by reference. These and numerous other negative mutations are often more deleterious than mutations causing the production of no gene product (null mutations or promoters that function in plant cells are known to those null alleles). For example, SEQ ID NO: 839-842 are con skilled in the art and available for use in recombinant poly structed to encodeagl11 protein with K-box deleted or C-ter nucleotides of the present invention to provide for expression minal domain deleted. AGL11 or MADS box protein can be of desired genes in transgenic plant cells. considered as having three functional domains. There is an 0034. In other aspects of the invention, preferential N-terminal DNA-binding domain (the MADS box), a more expression in plant green tissues is desired. Promoters of distal dimerization domain (the K-box) and a C-terminal interest for Such uses include those from genes such as Ara domain that is usually involved in interactions with other bidopsis thaliana ribulose-1,5-bisphosphate carboxylase proteins. In plants the region between the MADS box and the (Rubisco) small subunit (Fischhoff et al. (1992) Plant Mol K-box has been shown to be important for DNA binding in Biol. 20:81-93), aldolase and pyruvate orthophosphate diki some proteins and is often referred to as the 1-box (Fan et al., nase (PPDK) (Taniguchi et al. (2000) Plant Cell Physiol. 1997). Several different classes of dominant negative con 41 (11):42-48). structs are considered. Deletion or inactivation of the DNA 0035. Furthermore, the promoters may be altered to con binding domain can create proteins that are able to dimerize tain multiple "enhancer sequences' to assist in elevating gene with their native full length counterparts as well as other expression. Such enhancers are known in the art. By includ natural dimerization partners. Likewise, removal of the C-ter ing an enhancer sequence with Such constructs, the expres minal domain can allow dimerization with both the native sion of the selected protein may be enhanced. These enhanc protein and it’s natural dimerization partners. In both cases ers often are found 5' to the start of transcription in a promoter these types of constructs disable both the target protein and that functions in eukaryotic cells, but can often be inserted any other protein capable of interacting with the K-box. upstream (5') or downstream (3') to the coding sequence. In 0031. In other embodiments of the invention a constitu Some instances, these 5' enhancing elements are introns. Par tively active mutant is constructed to achieve the desired ticularly useful as enhancers are the 5' introns of the rice actin effect. For example, SEQID NO: 831 and SEQID NO: 832 1 (see U.S. Pat. No. 5,641,876) and rice actin 2 genes, the encode only the kinase domain from a calcium-dependent maize alcohol dehydrogenase gene intron, the maize heat protein kinase (CDPK). CDPK1 has a domain structure simi shock protein 70 gene intron (U.S. Pat. No. 5,593.874) and lar to other calcium-dependant protein kinases in which the the maize shrunken 1 gene. protein kinase domain is separated from four efhand domains 0036. In other aspects of the invention, sufficient expres by 42 amino acid 'spacer” region. Calcium-dependant pro sion in plant seed tissues is desired to effect improvements in tein kinases are thought to be activated by a calcium-induced seed composition. Exemplary promoters for use for seed conformational change that results in movement of an auto composition modification include promoters from seed genes inhibitory domain away from the protein kinase such as napin (U.S. Pat. No. 5,420,034), maize L3 oleosin (Yokokura et al., 1995). Thus, constitutively active proteins (U.S. Pat. No. 6,433,252), Zein Z27 (Russell et al. (1997) can be made by over expressing the protein kinase domain Transgenic Res. 6(2): 157-166), globulin 1 (Belanger et al alone. (1991) Genetics 129:863-872), glutelin 1 (Russell (1997) 0032 DNA constructs are assembled using methods well Supra), and peroxiredoxin antioxidant (Perl) (Stacy et al. known to persons of ordinary skill in the art and typically (1996) Plant Mol Biol. 31(6): 1205-1216). comprise a promoter operably linked to DNA, the expression 0037 Recombinant DNA constructs prepared in accor of which provides the enhanced agronomic trait. Other con dance with the invention will also generally include a 3' struct components may include additional regulatory ele element that typically contains a polyadenylation signal and ments, such as 5' leasders and introns for enhancing transcrip site. Well-known 3' elements include those from Agrobacte tion, 3' untranslated regions (such as polyadenylation signals rium tumefaciens genes Such as noS3", tml 3', tmr3', tmS3, ocs and sites), DNA for transit or signal peptides. 3', tr73', for example disclosed in U.S. Pat. No. 6,090,627, 0033 Numerous promoters that are active in plant cells incorporated herein by reference; 3' elements from plant have been described in the literature. These include promoters genes such as wheat (Triticum aesevitum) heat shock protein present in plant genomes as well as promoters from other 17 (Hsp173"), a wheat ubiquitin gene, a wheat fructose-1,6- Sources, including nopaline synthase (NOS) promoter and biphosphatase gene, a rice glutelin gene a rice lactate dehy octopine synthase (OCS) promoters carried on tumor-induc drogenase gene and a rice beta-tubulin gene, all of which are ing plasmids of Agrobacterium tumefaciens, caulimovirus disclosed in U.S. published patent application 2002/01928.13 promoters such as the cauliflower mosaic virus. For instance, A1, incorporated herein by reference; and the pea (Pisum see U.S. Pat. Nos. 5,858,742 and 5,322,938, which disclose sativum) ribulose biphosphate carboxylase gene (rbs 3), and versions of the constitutive promoter derived from cauli 3' elements from the genes within the host plant. US 2008/0148432 A1 Jun. 19, 2008

0038 Constructs and vectors may also include a transit which bind to the proteins disclosed herein. Means for pre peptide for targeting of a gene target to a plant organelle, paring and characterizing antibodies are well known in the art particularly to a chloroplast, leucoplast or other plastid (See, e.g., Antibodies: A Laboratory Manual, Cold Spring organelle. For descriptions of the use of chloroplast transit Harbor Laboratory, 1988; incorporated herein by reference). peptides see U.S. Pat. No. 5,188,642 and U.S. Pat. No. 5,728, The methods for generating monoclonal antibodies (mAbs) 925, incorporated herein by reference. For description of the generally begin along the same lines as those for preparing transit peptide region of an Arabidopsis EPSPS gene useful in polyclonal antibodies. Briefly, a polyclonal antibody is pre the present invention, see Klee, H. J. et al (MGG (1987) pared by immunizing an animal with an immunogenic com 210:437-442). position in accordance with the present invention and collect 0039 Transgenic plants comprising or derived from plant ing antisera from that immunized animal. A wide range of cells of this invention transformed with recombinant DNA animal species can be used for the production of antisera. can be further enhanced with stacked traits, e.g. a crop plant Typically the animal used for production of anti-antisera is a having an enhanced trait resulting from expression of DNA rabbit, a mouse, a rat, a hamster, a guinea pig or a goat. disclosed herein in combination with herbicide and/or pest Because of the relatively large blood volume of rabbits, a resistance traits. For example, genes of the current invention rabbit is a preferred choice for production of polyclonal anti can be stacked with other traits of agronomic interest, Such as bodies. a trait providing herbicide resistance, or insect resistance, Such as using a gene from Bacillus thuringensis to provide 0041 AS is well known in the art, a given composition may resistance against lepidopteran, coliopteran, homopteran, vary in its immunogenicity. It is often necessary therefore to hemiopteran, and other insects. Herbicides for which trans boost the host immune system, as may be achieved by cou genic plant tolerance has been demonstrated and the method pling a peptide or polypeptide immunogen to a carrier. Exem of the present invention can be applied include, but are not plary and preferred carriers are keyhole limpet hemocyanin limited to, glyphosate, dicamba, glufosinate, Sulfonylurea, (KLH) and bovine serum albumin (BSA). Other albumins bromoxynil and norflurazon herbicides. Polynucleotide mol Such as ovalbumin, mouse serum albumin or rabbit serum ecules encoding proteins involved in herbicide tolerance are albumin can also be used as carriers. Means for conjugating a well-known in the art and include, but are not limited to, a polypeptide to a carrier protein are well known in the art and polynucleotide molecule encoding 5-enolpyruvylshikimate include using glutaraldehyde, m-maleimidobencoyl-N-hy 3-phosphate synthase (EPSPS) disclosed in U.S. Pat. Nos. droxysuccinimide ester, carbodiimide and bis-biazotized 5,094,945;5,627,061; 5,633,435 and 6,040.497 for imparting benzidine. glyphosate tolerance; polynucleotide molecules encoding a 0042. As is also well known in the art, the immunogenicity glyphosate (GOX) disclosed in U.S. Pat. No. of a particular immunogen composition can be enhanced by 5,463,175 and a glyphosate-N-acetyl transferase (GAT) dis the use of non-specific stimulators of the immune response, closed in U.S. Patent Application publication 2003/0083480 known as adjuvants. Exemplary and preferred adjuvants A1 also for imparting glyphosate tolerance; dicamba include complete Freund's adjuvant (a non-specific stimula monooxygenase disclosed in U.S. Patent Application publi tor of the immune response containing killed Mycobacterium cation 2003/0135879 A1 for imparting dicamba tolerance; a tuberculosis), incomplete Freund's adjuvants and aluminum polynucleotide molecule encoding bromoxynil hydroxide adjuvant. (Bxn) disclosed in U.S. Pat. No. 4,810,648 for imparting 0043. The amount of immunogen composition used in the bromoxynil tolerance; a polynucleotide molecule encoding production of polyclonal antibodies varies upon the nature of phytoene desaturase (crtI) described in Misawa et al. (1993) the immunogen as well as the animal used for immunization. Plant J4:833-840 and Misawa etal, (1994) Plant J 6:481-489 A variety of routes can be used to administer the immunogen for norflurazon tolerance; a polynucleotide molecule encod (Subcutaneous, intramuscular, intradermal, intravenous and ing acetohydroxyacid synthase (AHAS, aka ALS) described intraperitoneal). The production of polyclonal antibodies in Sathasiivanet al. (1990) Nucl. Acids Res. 18:2188-2193 for may be monitored by sampling blood of the immunized ani imparting tolerance to Sulfonylurea herbicides; polynucle mal at various points following immunization. A second, otide molecules known as bar genes disclosed in DeBlock, et booster, injection may also be given. The process of boosting al. (1987) EMBO.J. 6:2513-2519 for imparting glufosinate and titering is repeated until a suitable titer is achieved. When and bialaphos tolerance; polynucleotide molecules disclosed a desired level of immunogenicity is obtained, the immunized in U.S. Patent Application Publication 2003/010609 A1 for animal can be bled and the serum isolated and stored, and/or imparting N-amino methyl phosphonic acid tolerance; poly the animal can be used to generate mabs. nucleotide molecules disclosed in U.S. Pat. No. 6,107,549 for 0044 mAbs may be readily prepared through use of well imparting pyridine herbicide resistance; molecules and meth known techniques, such as those exemplified in U.S. Pat. No. ods for imparting tolerance to multiple herbicides Such as 4,196,265, incorporated herein by reference. Typically, this glyphosate, atrazine, ALS inhibitors, isoxoflutole and glufo technique involves immunizing a suitable animal with a sinate herbicides are disclosed in U.S. Pat. No. 6,376,754 and selected immunogen composition, e.g., a purified or partially U.S. Patent Application Publication 2002/0112260, all of purified antifungal protein, polypeptide or peptide. The said U.S. patents and patent application Publications are immunizing composition is administered in a manner effec incorporated herein by reference. Molecules and methods for tive to stimulate antibody producing cells. Rodents such as imparting insect/nematode?virus resistance is disclosed in mice and rats are preferred animals, however, the use of U.S. Pat. Nos. 5,250,515; 5,880,275; 6,506,599; 5,986,175 rabbit, sheep, or frog cells is also possible. The use of rats may and U.S. Patent Application Publication 2003/015.0017 A1, provide certain advantages (Goding, 1986, pp. 60-61), but all of which are incorporated herein by reference. mice are preferred, with the BALB/c mouse being most pre 0040. In particular embodiments, the inventors contem ferred as this is most routinely used and generally gives a plate the use of antibodies, either monoclonal or polyclonal higher percentage of stable fusions. US 2008/0148432 A1 Jun. 19, 2008

0045. Following immunization, somatic cells with the of nucleotides (HAT medium). Where azaserine is used, the potential for producing antibodies, specifically B lympho media is Supplemented with hypoxanthine. cytes (B cells), are selected for use in the mAb generating 0051. The preferred selection medium is HAT. Only cells protocol. These cells may be obtained from biopsied spleens, capable of operating nucleotide Salvage pathways are able to tonsils or lymph nodes, or from a peripheral blood sample. survive in HAT medium. The myeloma cells are defective in Spleen cells and peripheral blood cells are preferred, the key of the Salvage pathway, e.g., hypoxanthine former because they are a rich Source of antibody-producing phosphoribosyltransferase (HPRT), and they cannot survive. cells that are in the dividing plasmablast stage, and the latter The B-cells can operate this pathway, but they have a limited because peripheral blood is easily accessible. Often, a panel life span in culture and generally die within about two weeks. of animals will have been immunized and the spleen of ani Therefore, the only cells that can survive in the selective mal with the highest antibody titer will be removed and the media are those hybrids formed from myeloma and B-cells. spleen lymphocytes obtained by homogenizing the spleen 0052. This culturing provides a population of hybridomas with a syringe. Typically, a spleen from an immunized mouse from which specific hybridomas are selected. Typically, contains approximately 5x10 to 2x10 lymphocytes. selection of hybridomas is performed by culturing the cells by 0046. The antibody-producing B lymphocytes from the single-clone dilution in microtiter plates, followed by testing immunized animal are then fused with cells of an immortal the individual clonal supernatants (after about two to three myeloma cell, generally one of the same species as the animal weeks) for the desired reactivity. The assay should be sensi that was immunized. Myeloma cell lines suited for use in tive, simple and rapid, Such as radioimmunoassays, enzyme hybridoma-producing fusion procedures preferably are non immunoassays, cytotoxicity assays, plaque assays, dot antibody-producing, have high fusion efficiency, and enzyme immunobinding assays, and the like. deficiencies that render them incapable of growing in certain 0053. The selected hybridomas would then be serially selective media which support the growth of only the desired diluted and cloned into individual antibody-producing cell fused cells (hybridomas). lines, which clones can then be propagated indefinitely to 0047 Any one of a number of myeloma cells may be used, provide mabs. The cell lines may be exploited for mAb as are known to those of skill in the art (Goding, 1986, pp. production in two basic ways. A sample of the hybridoma can 65-66; Campbell, 1984, pp. 75-83). For example, where the be injected (often into the peritoneal cavity) into a histocom immunized animal is a mouse, one may use P3-X63/Ag8, patible animal of the type that was used to provide the somatic X63-Ag8.653, NS1/1. Ag 41, Sp210-Ag14, FO, NSO/U, and myeloma cells for the original fusion. The injected ani MPC-11, MPC11-X45-GTG 1.7 and S194/5XX0 Bul; for mal develops tumors secreting the specific monoclonal anti rats, one may use R210.RCY3, Y3-Ag 1.2.3, IR983F and body produced by the fused cell hybrid. The body fluids of the 4B210; and U-266, GM1500-GRG2, LICR-LON-HMy2 and animal. Such as serum or ascites fluid, can then be tapped to UC729-6 are all useful in connection with humancell fusions. provide mabs in high concentration. The individual cell lines 0048 One preferred murine myeloma cell is the NS-1 could also be cultured in vitro, where the mabs are naturally myeloma cell line (also termed P3-NS-1-Ag4-1), which is secreted into the culture medium from which they can be readily available from the NIGMS Human Genetic Mutant readily obtained in high concentrations. mAbs produced by Cell Repository by requesting cell line repository number either means may be further purified, if desired, using filtra GM3573. Another mouse myeloma cell line that may be used tion, centrifugation and various chromatographic methods is the 8-azaguanine-resistant mouse murine myeloma SP2/0 such as HPLC or affinity chromatography. non-producer cell line. 0049 Methods for generating hybrids of antibody-pro Plant Cell Transformation Methods ducing spleen or lymph node cells and myeloma cells usually 0054 Numerous methods for transforming plant cells comprise mixing somatic cells with myeloma cells in a 2:1 with recombinant DNA are known in the art and may be used ratio, though the ratio may vary from about 20:1 to about 1:1, in the present invention. Two commonly used methods for respectively, in the presence of an agent or agents (chemical plant transformation are Agrobacterium-mediated transfor or electrical) that promote the fusion of cell membranes. mation and microprojectile bombardment. Microprojectile Fusion methods using Spend virus have been described bombardment methods are illustrated in U.S. Pat. Nos. 5,015, (Kohler and Milstein, 1975; 1976), and those using polyeth 580 (soybean): 5,550,318 (corn); 5,538,880 (corn); 5,914, ylene glycol (PEG), such as 37% (v/v) PEG, (Gefter et al., 451 (soybean); 6,160,208 (corn); 6,399,861 (corn)and 6,153, 1977). The use of electrically induced fusion methods is also 812 (wheat) and Agrobacterium-mediated transformation is appropriate (Goding, 1986, pp. 71-74). described in U.S. Pat. Nos. 5,159,135 (cotton); 5,824,877 0050 Fusion procedures usually produce viable hybrids at (soybean); 5,591,616 (corn); and 6,384.301 (soybean), all of low frequencies, about 1x10 to 1x10. However, this does which are incorporated herein by reference. For Agrobacte not pose a problem, as the viable, fused hybrids are differen rium tumefaciens based plant transformation system, addi tiated from the parental, unfused cells (particularly the tional elements present on transformation constructs will unfused myeloma cells that would normally continue to include T-DNA left and right border sequences to facilitate divide indefinitely) by culturing in a selective medium. The incorporation of the recombinant polynucleotide into the selective medium is generally one that contains an agent that plant genome. blocks the de novo synthesis of nucleotides in the tissue 0055. In general it is useful to introduce recombinant DNA culture media. Exemplary and preferred agents are aminop randomly, i.e. at a non-specific location, in the genome of a terin, methotrexate, and azaserine. Aminopterin and methotr target plant line. In special cases it may be useful to target exate block de novo synthesis of both purines and pyrim recombinant DNA insertion in order to achieve site-specific idines, whereas azasenne blocks only purine synthesis. integration, for example to replace an existing gene in the Where aminopterin or methotrexate is used, the media is genome, to use an existing promoter in the plant genome, or Supplemented with hypoxanthine and thymidine as a source to inserta recombinant polynucleotide at a predetermined site US 2008/0148432 A1 Jun. 19, 2008 known to be active for gene expression. Several site specific that are stably transformed by receiving and integrating a recombination systems exist which are known to function transgenic DNA construct into their genomes. Preferred implants include cre-lox as disclosed in U.S. Pat. No. 4,959, marker genes provide selective markers which confer resis 317 and FLP-FRT as disclosed in U.S. Pat. No. 5,527,695, tance to a selective agent, such as an antibiotic or herbicide. both incorporated herein by reference. Any of the herbicides to which plants of this invention may be 0056 Transformation methods of this invention are pref resistant are useful agents for selective markers. Potentially erably practiced in tissue culture on media and in a controlled transformed cells are exposed to the selective agent. In the environment. “Media' refers to the numerous nutrient mix population of Surviving cells will be those cells where, gen tures that are used to grow cells in vitro, that is, outside of the erally, the resistance-conferring gene is integrated and intact living organism. Recipient cell targets include, but are expressed at sufficient levels to permit cell survival. Cells not limited to, meristem cells, callus, immature embryos and may be tested further to confirm stable integration of the gametic cells such as microspores, pollen, sperm and egg exogenous DNA. Commonly used selective marker genes cells. It is contemplated that any cell from which a fertile plant include those conferring resistance to antibiotics such as may be regenerated is useful as a recipient cell. Callus may be kanamycin and paromomycin (nptII), hygromycin B (aph IV) initiated from tissue sources including, but not limited to, and gentamycin (aac3 and aacC4) or resistance to herbicides immature embryos, seedling apical meristems, microspores Such as glufosinate (bar or pat) and glyphosate (aroA or and the like. Cells capable of proliferating as callus are also EPSPS). Examples of such selectable are illustrated in U.S. recipient cells for genetic transformation. Practical transfor Pat. Nos. 5,550,318; 5,633,435; 5,780,708 and 6,118,047, all mation methods and materials for making transgenic plants of of which are incorporated herein by reference. Selectable this invention, for example various media and recipient target markers which provide an ability to visually identify trans cells, transformation of immature embryo cells and Subse formants can also be employed, for example, a gene express quent regeneration offertile transgenic plants are disclosed in ing a colored or fluorescent protein such as a luciferase or U.S. Pat. Nos. 6,194.636 and 6.232,526, which are incorpo green fluorescent protein (GFP) or a gene expressing a beta rated herein by reference. glucuronidase or uidA gene (GUS) for which various chro 0057 The seeds of transgenic plants can be harvested mogenic Substrates are known. from fertile transgenic plants and be used to grow progeny 0059) Plant cells that survive exposure to the selective generations of transformed plants of this invention including agent, or plant cells that have been scored positive in a screen hybrid plants line for selection of plants having an enhanced ing assay, may be cultured in regeneration media and allowed trait. In addition to direct transformation of a plant with a recombinant DNA, transgenic plants can be prepared by to mature into plants. Developing plantlets regenerated from crossing a first plant having a recombinant DNA with a sec transformed plant cells can be transferred to plant growth ond plant lacking the DNA. For example, recombinant DNA mix, and hardened off for example, in an environmentally can be introduced into first plant line that is amenable to controlled chamber at about 85% relative humidity, 600 ppm transformation to produce a transgenic plant which can be CO, and 25-250 microeinsteins mis' of light, prior to crossed with a second plant line to introgress the recombinant transfer to a greenhouse or growth chamber for maturation. DNA into the second plant line. A transgenic plant with Plants are regenerated from about 6 weeks to 10 months after recombinant DNA providing an enhanced trait, e.g. enhanced a transformant is identified, depending on the initial tissue. yield, can be crossed with transgenic plant line having other Plants may be pollinated using conventional plant breeding recombinant DNA that confers another trait, for example methods known to those of skill in the art and seed produced, herbicide resistance or pest resistance, to produce progeny for example self-pollination is commonly used with trans plants having recombinant DNA that confers both traits. genic corn. The regenerated transformed plant or its progeny Typically, in Such breeding for combining traits the trans seed or plants can be tested for expression of the recombinant genic plant donating the additional trait is a male line and the DNA and selected for the presence of enhanced agronomic transgenic plant carrying the base traits is the female line. The trait. progeny of this cross will segregate Such that some of the plants will carry the DNA for both parental traits and some Transgenic Plants and Seeds will carry DNA for one parental trait; such plants can be identified by markers associated with parental recombinant 0060 Transgenic plants derived from the plant cells of this DNA, e.g. marker identification by analysis for recombinant invention are grown to generate transgenic plants having an DNA or, in the case where a selectable marker is linked to the enhanced trait as compared to a control plant and produce recombinant, by application of the selecting agent Such as a transgenic seed and haploid pollen of this invention. Such herbicide for use with a herbicide tolerance marker, or by plants with enhanced traits are identified by selection of trans selection for the enhanced trait. Progeny plants carrying DNA formed plants or progeny seed for the enhanced trait. For for both parental traits can be crossed back into the female efficiency a selection method is designed to evaluate multiple parent line multiple times, for example usually 6 to 8 genera transgenic plants (events) comprising the recombinant DNA, tions, to produce a progeny plant with Substantially the same for example multiple plants from 2 to 20 or more transgenic genotype as one original transgenic parental line but for the events. Transgenic plants grown from transgenic seed pro recombinant DNA of the other transgenic parental line vided herein demonstrate improved agronomic traits that 0058. In the practice of transformation DNA is typically contribute to increased yield or other trait that provides introduced into only a small percentage of target plant cells in increased plant value, including, for example, improved seed any one transformation experiment. Marker genes are used to quality. Of particular interest are plants having enhanced provide an efficient system for identification of those cells water use efficiency, enhanced cold tolerance, increased US 2008/0148432 A1 Jun. 19, 2008 9 yield, enhanced nitrogen use efficiency, enhanced seed pro- “NUCSEQ which identifies a DNA sequence from SEQID tein and enhanced seed oil. NO: 1 to 741. 0061 Table 1 provides a list of protein encoding DNA “Base Vector” is a reference to the identifying number in (“genes”) that are useful as recombinant DNA for production Table 4 of base vectors used for construction of the transfor of transgenic plants with enhanced agronomic trait, the ele- mation vectors of the recombinant DNA. Construction of ments of Table 1 are described by reference to: plant transformation constructs is illustrated in Example 1. “PEP SEQ which identifies an amino acid sequence from “PROTEIN NAME’ which is a common name for protein SEQID NO: 742 to 1482. encoded by the recombinant DNA.

TABLE 1.

PEP NUC SEQ SEQ ID ID Base NO NO GENEID Vector PPROTEIN NAME 742 1 PHEOOO1089 1179 Arabidopsis NAC domain transcription factor 743 2 PHEOOO1133 1223 yeast aspartate aminotransferase (AAT2) 744 3 PHEOOO1134 1224 yeast aspartate aminotransferase (AAT1)- CAA97SSO 745 4 PHEOOO1135 1225 E. coli aspC-1651442 746 5 PHEOOO1181. 1271 rice IAA1-like 1-AJ251791 747 6 PHEOOO1227 1317 yeast CTT1-NP 011602 748 7 PHEOOO1228 1318 yeast STE20-AAB69747 749 8 PHEOOO1247 1338 Arabidopsis cyclin D3-like-CGPG 710 750 9 PHEOOO1252. 1343 Arabidopsis arabinogalactan-protein 1 751 10 PHEOOO1267 1358 Arabidopsis hypothetical protein 752 11 PHEOOO1275. 1336 Arabidopsis CIP8 753 12 PHEOOO1287 1377 Arabidopsis hypothetical protein 754 13 PHEOOO1382. 1474 rice Short-Root SHR-like transcriptional factor 1 sequence 755 14 PHEOOO1531. 1622 Bacilius subtilis GDH 756 15 PHEOOO1532 1623 Saccharomyces cerevisiae GDH-1431821 757 16 PHEOOO1535. 1626 Pseudomonas fluorescens GDH 758 17 PHEOOO1536. 1627 Bacilius halodurans GDH 759 18 PHEOOO1538 1629 Synechocystis sp. 6803 GDH 760 19 PHEOOO1593. 1699 5 yeast transketolase 761 20 PHEOOO1593. 1700 yeast transketolase 762 21 PHEOOO2O70 218O rice G664-like 2 763 22 PHEOOO2128 2236 E. coli phosphoenolpyruvate carboxylase 764 23 PHEOOO2258 69 2 corn inphI-S851D 765 24 PHEOOO2259 68 2 corn inphI-S849D 766 25 PHEOOO2260 70 2 corn mph.I-S849D-S851D 767 26 PHEOOO2645 2764 Arabidopsis GH3 protein 3 768 27 PHEOOO2976 3126 Yeast Serine. Threonine Protein Kinase 769 28 PHEOOO2999 3149 Yeast Stereospecific (2R,3R)-2,3-butanediol dehydrogenase with similarity to alcoholsorbitol dehydrogenases 770 29 PHEOOO3OOO 3150 Yeast Car2p: Ornithine aminotransferase 771 30 PHEOOO3OO1 3151 Yeast alpha-Mannosidase 772 31 PHEOOO3OO2 3152 Yeast Putative indole-3-pyruvate decarboxylase 773 32 PHEOOO3004 3154 Yeast Citrate synthase 774 33 PHEOOO3006. 31.56 Yeast Galactose-induced protein with strong similarity to crystallin protein of vertebrate eye lens 775 34 PHEOOO3007 3157 Yeast Phosphoglycerate mutase 776 35 PHEOOO3008 3158 Yeast Heat shock protein of 26 kDa, expressed during entry to stationary phase and induced by osmostress 777 36 PHEOOO3O12 3162 Yeast Neutral trehalase (alpha, alpha trehalase), catalyzes the conversion of intracellular trehalose to glucose 778 37 PHEOOO3O14 3164. YEAST P16547 MITOCHONDRIAL OUTER MEMBRAINE 4S KD PROTEIN 779 38 PHEOOO3O22 3172 Yeast Protein of unknown function, has strong similarity to Tallip (Transaldolase PFAM domain) 78O 39 PHEOOO3O31 3181 Yeast Long-chain fatty acid CoA (fatty acid activator 1) 781 40 PHEOOO3035 3.185 Yeast Glucokinase, specific for aldohexoses 782 41 PHEOOO3O37 31.87 Yeast Aldose reductase with NADPH specificity; induced by osmotic stress 783 42 PHEOOO3O44 3.194 Yeast Calcium/calmodulin-dependent serine/threonine protein kinase (CaM kinase) type II US 2008/0148432 A1 Jun. 19, 2008 10

TABLE 1-continued

PEP NUC SEQ SEQ ID ID Base NO NO GENEID Vector PPROTEIN NAME 784 43 PHEOOO3056 3206 1 yeast CBK1 785 44 PHEOOO3166 3368 6 Agrobacterium tumefaciens str. C58 sucrose phosphorylase 786 45 PHEOOO3191, 3390 rice G1225-like 1 787 46 PHEOOO3266 3485 Arabidopsis unknown protein 788 47 PHEOOO3295 3515 rice unknown protein 789 48 PHEOOO3358. 3581 rice dwarfa-like 790 49 PHEOOO3416 3656 Arabidopsis homogentisate phytylprenyltransferase 791 50 PHEOOO3457 3688 rice G1660 like 1 792 51 PHEOOO35O2 3748 rice G.2239 like1 793 52 PHEOOO3506 3752 rice G2317 like2 794 53 PHEOOO3522 3768 rice G2536 like1 795 54 PHEOOO3592 3838 rice G864 like1 796 55 PHEOOO3595 3841 rice G46 like1 797 56 PHEO003633 3891 rice G293.0 like 1 798 57 PHEOOO3635. 3893 rice G2969 like2 799 58 PHEOOO3663 3921 rice G1108 like2 800 59 PHEOOO367O 3928 rice G1792 like3 8O1 60 PHEOOO3672 3930 rice G1013 like2 802 61 PHEOOO3675 3933 rice G1493 like3 803 62 PHEOOO3945. 4522 Arabidopsis CGPG2571 804 63 PHEOOO1066 1156 yeast SIP1-AAB64887 805 64 PHEOOO1471. 1563 rice NH4-uniport AMT1-like sequence 806 65 PHEOOO1530 1621 Emericeia nidulians GDH 807 66 PHEOOO2940 3090 Yeast Proline oxidase (proline dehydrogenase) 808 67 PHEOOO2957. 3107 Yeast MIP (member of major intrinsic protein family of transmembrane channels) 809 68 PHEOOO2961: 3111 Yeast protein with 3 RRM domains (RNA recongnition motifs) 810 69 PHEOOO2962. 3112 Yeast Putative Dual Specificity Phosphatase 811 70 PHEOOO3003 3153 Yeast Aromatic amino acid aminotransferase II 812 71 PHEOOO3O24 3174 Yeast protein of unknown function 813 72 PHE0003042. 3192 Yeast Possible 6-phosphogluconolactonase 814 73 PHEOOO3236. 3451 Arabidopsis unknown protein 815 74 PHEOOO328O 3499 Arabidopsis DUF6 816 75 PHEOOOOO35 61 corn lip 19 817 76 PHEOOOOO36 62 corn EF1-alpha 818 77 PHEOOOO130 220 b-glucosidase-aggregating factor 819 78 PHEOOOO134 224 LEA protein EMB5 82O 79 PHEOOOO135 225 821 80 PHEOOOO136 226 822 81 PHEOOOO139 229 ascorbate peroxidase 823 82 PHE0000141. 231 cp drought-induced stress protein 824 83 PHE0000142. 232 2 rab7a 825 84 PHE0000146 236 rab11d 826 85 PHEOOOO148 238 protein kinase 827 86 PHEOOOO149 239 2 EREbp-like AP2 domain TF 828 87 PHEOOOO150 240 protein kinase 829 88 PHEOOOO189 282 soy HSP20 (HS11) 830 89 PHEOOOO2OO 293 protein phosphatase 2C-like 831 90 PHE0000211. 304 2 CDPK kinase domain 832 91 PHEOOOO213 306 CDPK kinase domain 833 92 PHEOOOO368 459 rice millip19 834 93 PHEOOOO369 460 OSKS 835 94 PHEOOOO370 461 abscisic acid-inducible protein kinase 836 95 PHEOOOO780 853 Arabidopsis ag|11 837 96 PHEOOOO782. 855 corn agl11-like 1 838 97 PHEOOOO783 856 rice MADS3-L37528 839 98 PHEOOOO785 858 Arabidopsis ag|11 delta C-terminus 840 99 PHEOOOO791 864 Soyagl11-like 1 delta C-terminus 841 100 PHEOOOO792 865 soy ag|11-like 1 delta K-box 842 101 PHE0000794. 867 rice MADS3 delta C-terminus-L37528 843 102 PHEOOOO821. 896 corn dufó 10 844 103 PHEOOO1051 1141 corn Zinc finger protein 845 104 PHEOOO1053 4292 corn NAM-like protein 846 105 PHEOOO1054 1144. corn QKI-like RNA-binding protein 847 106 PHEOOO1055 1145 corn putative kinase regulatory Subunit 848 107 PHEOOO1056 1146 corn unknown protein US 2008/0148432 A1 Jun. 19, 2008 11

TABLE 1-continued

PEP NUC SEQ SEQ ID ID Base NO NO GENEID Vector PPROTEIN NAME 849 108 PHEOOO1057 47 corn thionin-like protease inhibitor 850 109 PHEOOO1064 S4 yeast GAL83-Q04739 851 O PHEOOO1071 61 Arabidopsis AP2 domain transcription factor 852 1 PHEOOO1073 63 Arabidopsis myb domain transcription factor 853 2 PHEOOO1075 65 Arabidopsis myb domain transcription factor 854 3 PHEOOO1077 67 Arabidopsis GARP domain transcription factor 855 4 PHEOOO1083 73 Arabidopsis AP2 domain transcription factor 856 5 PHEOOO1095 85 Arabidopsis bZIP domain transcription factor 857 6 PHEOOO1096 86 Arabidopsis bZIP domain transcription factor 858 7 PHEOOO1097 87 Arabidopsis bZIPTGA domain transcription factor 859 8 PHEOOO1098 88 Arabidopsis GATA domain transcription factor 860 9 PHEOOO1099 89 Arabidopsis bZIP domain transcription factor 861 120 PHEOOO1101 91 Arabidopsis GATA domain transcription factor 862. 121 PHEOOO1107 97 rice CCA1-like 1 863 122 PHEOOO1108 98 rice alanine aminotransferase 2-AAK52114 864. 123 PHEOOO1109 99 corn alanine aminotransferase 1 865 124 PHEOOO1110 200 corn alanine aminotransferase 4 866 125 PHEOOO1115 205 yeast 5-aminolevulinic acid synthase mature form-PO99SO 867 126 PHEOOO1120 210 corn G1820-like 1 868 127 PHEOOO1123 213 corn G1820-like 2 869 128 PHEOOO1125 215 soy G1820 like 2 870 129 PHEOOO1126 216 corn putative myb-related transcription factor 871. 130 PHEOOO1127 217 corn putative dehydration-responsive protein RD22 precursor 872 131 PHEOOO1128 218 corn homeodomain leucine Zipper protein 873 132 PHEOOO1130 220 corn acetohydroxyacid reductoisomerase, chloroplast precursor 874 133 PHEOOO1132 222 corn cytosolic aspartate transaminase 875 134 PHEOOO1138 228 corn hypothetical protein 876 135 PHEOOO1145 235 corn unknown protein 877 136 PHEOOO1146 236 corn MAP kinase kinase 878 137 PHEOOO1147 237 corn hypothetical protein 879 138 PHEOOO1148 238 corn transfactor-like protein 880 139 PHEOOO1151 241 corn dehydration-responsive protein RD22 precursor 881. 140 PHEOOO1152 242 corn histone H3 882 141 PHEOOO1153 243 corn calcium-dependent protein kinase 883 142 PHEOOO1154 244 corn histone H2B 884. 143 PHEOOO1156 246 corn ATFP4-like 885 144 PHEOOO1157 247 corn unknown protein 886 145 PHEOOO1158 248 corn glucose-6-phosphate:phosphate translocator precursor 887 146 PHEOOO1159 249 corn DnaJ-like protein 888 147 PHEOOO1167 257 rem1-Mu 889 148 PHEOOO1171 261 soy TU8-like 1 890 149 PHEOOO1172 262 soy TU8-like protein 2 891. 150 PHEOOO1179 269 corn AXR2-like 2 892 151 PHEOOO1198 288 soybean G175-like 2 893 152 PHEOOO1199 289 soybean G175-like 1 894 153 PHEOOO1208 298 soybean G.22-like 1 895 154 PHEOOO1211 301 soybean G867-like 1 896 155 PHEOOO1212 3O2 STE11-NP 013466 897 156 PHEOOO1214 304 soybean STE11-like 1 898 157 PHEOOO1215 305 soybean G1836-like 1 899 158 PHEOOO1220 310 soybean AtGSK3-like 2 900 159 PHEOOO1221 311 maize AtGSK3-like 1 901 160 PHEOOO1225 315 maize catalase-like 1 902 161 PHEOOO1238 328 maize cup1a-like 1 903 162 PHEOOO1239 329 maize AtGSK3-like 2 904 163 PHEOOO1241 331 Putative RNA Binding protein 905 164 PHEOOO1242 332 Protein Disulfide 906 165 PHEOOO1244 334 LIM DOmain Transcription Factor SF3 907 166 PHEOOO1245 335 Unkown Protein 908 167 PHEOOO1246 337 soy CIP8-like 1 909 168 PHEOOO1254 345 Arabidopsis hypothetical protein NM 111482 US 2008/0148432 A1 Jun. 19, 2008 12

TABLE 1-continued

PEP NUC SEQ SEQ ID ID Base NO NO GENEID Vector PPROTEIN NAME 910 169 PHEOOO1258 349 corn NADH:ubiquinone oxidoreductase 911 170 PHEOOO1259 350 soy NADH:ubiquinone oxidoreductase 912 171 PHEOOO1279 369 corn cysteine proteinase inhibitor 913 172 PHEOOO1280 370 soy cysteine proteinase inhibitor 914 173 PHEOOO1282 372 corn fatty acid elongase 1 915 174 PHEOOO1284 374 soy ADP-ribosylation factor 1 GTPase activating protein 916 175 PHEOOO1285 375 corn ADP-ribosylation factor 1 GTPase activating protein 917, 176 PHEOOO1291 381 soy hypothetical protein 918 177 PHEOOO1292 382 corn hypothetical protein 919 178 PHEOOO1297 387 corn cryptochrome like protein 1 920, 179 PHEOOO1299 389 corn cryptochrome like protein 5 921 180 PHEOOO1303 393 rice Pra2-like protein 2 922, 181 PHEOOO1306 396 soy Pra2-like protein 1 923 182 PHEOOO1312 402 corn CGPG 1145-like protein 1 924, 183 PHEOOO1313 403 corn CGPG 1145-like protein 2 925 184 PHEOOO1314 404 corn CGPG 1145-like protein 3 926 185 PHEOOO1332 423 rice SNF1-like protein 4-BAB61199 927 186 PHEOOO1333 424 rice SNF1-like protein 5-BAA96628 928, 187 PHEOOO1334 425 rice SNF1-like protein 6 OSK1-D82039 929 188 PHEOOO1337 428 rice AKIN-like protein 1 930 189 PHEOOO1339 430 soy AKIN-like protein 1 931 190 PHEOOO1343 434 soy SNF1-like protein 3 932 191 PHEOOO1346 437 corn SNF1-like protein 1 933 192 PHEOOO1347 438 corn SNF1-like protein 2 934 193 PHEOOO1349 440 corn SNF1-like protein 4 935 194 PHEOOO1350 441 corn SNF1-like1 protein 5 936 195 PHEOOO1354 445 corn SNF1-like1 protein 9 937 196 PHEOOO1355 446 corn SNF1-like1 protein 10 938 197 PHEOOO1356 447 corn SNF1-like1 protein 11 939 198 PHEOOO1357 448 corn SNF1-like protein 13 940 199 PHEOOO1358 449 corn SNF1-like protein 14 941 200 PHEOOO1359 450 corn SNF1-like protein 15 942 201 PHEOOO1360 451 Corn Putative CCR4 Associated Factor 943 202 PHEOOO1361 452 Soy Putative AP2 Domain Transcription Factor 944 203 PHEOOO1362 453 Soy GATA Binding Transcription Factor 945 204 PHEOOO1363 455 NAC1 type transcriptional activator 946 205 PHEOOO1364 456 Corn Putative Transcriptional Regulator X2 947 206 PHEOOO1375 467 Corn PRL 1 948. 207 PHEOOO1377 469 rice PINOID-like protein kinase 1 sequence 949 208 PHEOOO1380 472 rice PP2A regulatory subunit ARCN1-like 1 sequence 950 209 PHEOOO1385 477 maize IAA-alanine resistance protein IAR1 like sequence 951 210 PHEOOO1386 478 maize IAA-Ala hydrolase like 1 sequence 952 211 PHEOOO1389 481 maize nitrilase 1 like 1 sequence 953 212 PHEOOO1392 484 soybean root-specific kinase ARSK1 like SeleCo 954 213 PHEOOO1394 486 soybean MADS-box protein AGL14-like SeleCo 955 214 PHEOOO1397 489 soybean Ran binding protein RanBP1-like 1 SeleCo 956 2.15 PHEOOO1398 490 maize Ran binding protein RanBP1-like 1 SeleCo 957 216 PHEOOO1405 497 niftJ like protein Homolog 958 217 PHEOOO1411 503 photosystem-IPSI-F subunit precursor 959 218 PHEOOO1416 SO8 growth-on protein GRO10 96.O 219 PHEOOO1417 509 Transcription Factor Homolog BTF3 961 220 PHEOOO1418 510 Putative CGI-19 Protein Homolog 962 221 PHEOOO1421 513 glutamate dehydrogenase 963 222 PHEOOO1426 S18 Rice GATA Factor Homolog 964 223 PHEOOO1436 528 Methionine Synthase 965 224 PHEOOO1437 529 Putative ABC Transporter 966 225 PHEOOO1442 534 corn C-24 sterol methyltransferase 967 226 PHEOOO1444 536 soy hypothetical protein 96.8 227 PHEOOO1452 544 maize root hair Ted2-like sequence 969 228 PHEOOO1463 555 soybean homeodomain protein (GLABRA2) like sequence 970 229 PHEOOO1477 569 soybean werewolf (WER) like sequence US 2008/0148432 A1 Jun. 19, 2008 13

TABLE 1-continued

PEP NUC SEQ SEQ ID ID Base NO NO GENEID Vec or PPROTEIN NAME 971 230 PHEOOO1481 573 rice ethylene-response ETR1 like sequence 972 231 PHEOOO1483 575 Soybean xyloglucan endotransglycosylase (XET) like 1 sequence 973 232 PHEOOO1516 607 Zea Mays SelR Domain protein 974 233 PHEOOO1522 613 corn LPE1-like permease 6 975 234 PHEOOO1539 630 Soyglutamate dehydrogenase 976 235 PHEOOO1577 676 Rice Homolog Putative Transcription Factor X2 977 236 PHEOOO1602 713 rice rac-like GTP binding protein like 1 Sequence 978 237 PHEOOO1604 715 rice rac-like GTP binding protein like 3 Sequence 979 238 PHEOOO1605 716 rice rac-like GTP binding protein like 4 Sequence 98O 239 PHEOOO1606 717 rice rac-like GTP binding protein like 5 Sequence 981 240 PHEOOO1610 721 maize translation initiation factor 3 delta Subunit like sequence 982 241 PHEOOO1617 728 maize metal transporter ZIP8 like 1 sequence 983 242 PHEOOO1624 735 maize magnesium transporter, mrs2-2-like 1 Sequence 984 243 PHEOOO1629 740 maize low temperature and salt responsive protein LT16A-like 1 sequence 985 244 PHEOOO1642 753 rice salt-induced-zinc-finger-protein 1 Sequence 986 245 PHEOOO1649 760 Soybean salt-tolerance protein 1 Seqeunce 987 246 PHEOOO2017 2 28 Corn RING Finger Transcription Factor 988 247 PHEOOO2O232 34 Corn RING Finger Transcription Factor II 989 248 PHEOOO2O24 2 35 Corn RING Finger Transcription Factor III 990 249 PHEOOO2O27 2 38 Glutamine Synthetase 991 250 PHEO002041 2 51 Cornubiquitin fusion proteinribosomal protein S27a 992 251 PHEO002042 2 52 soybean AthSP17.6A like 1 sequence 993 252 PHEO002049 2 59 Ribulose bisphosphate carboxylase small chain chloroplast precursor 994 253 PHEO002052 2 62 Corn ligand-gated channel-like protein precursor 995 254 PHEO002054 2 64 Corn Nitrilase 1 996 255 PHEOOO2055 2 65 Corn Tic 22 Like protein 997 256 PHEOOO2O67 2 77 corn G664-like 5 998 257 PHEOOO2O68 2 78 corn G664-like 6 999 258 PHEOOO2O71 2 81 soy G664-like 2 OOO 259 PHEOOO2080 4290 corn Sec61 OO1 260 PHEO002081 2 91 soy seco1 OO2 261 PHEOOO2O88 2 98 rice osr8 OO3 262 PHEOOO2095 2 205 Corn Calmodulin EF hand OO)4 263 PHEOOO2.096 2 2O6 Rice Calmodulin EF Hand 005 264 PHEOOO2099 2 209 Rice Putative Calmodulin EF Hand Protein OO6 265 PHEOOO2103 2 213 rice Bobs kinase 2 OO7 266 PHEO002123 2 231 4 rice phosphoenolpyruvate carboxylase 2 O08 267 PHEOOO2125 2 233 corn phosphoenolpyruvate carboxylase 2 O09 268 PHEOOO2127 2 235 soy phosphoenolpyruvate carboxylase 2 O10 269 PHEOOO2145 2 253 Corn Unkown Protein O11 270 PHEOOO2148 2 256 Corn Cytochrome P450 O12 271 PHEOOO2163 2 270 Corn Protein (Similar to peptidyl prolyl isomerase) 272 PHEOOO2164 2 271 Corn Protein (similar to yippie-1 & a peptidyl prolyl isomerase) O14 273 PHEOOO2183. 2 290 soybean Hsp17.7 like 1 sequence O15 274 PHEOOO2189 2 296 maize pyrroline-5-carboxylate synthetase like 2 sequence O16 275 PHEOOO2202 2 309 soy annexin 1 O17 276 PHEOOO2203 2 310 soy annexin 3 O18 277 PHEOOO2204 2 311 soy annexin 4 O19 278 PHEOOO2210 2 317 soybean drought-induced protein BnD22 like 1 sequence O20 279 PHEOOO2224 2 331 maize drought-induced protein Dil 9-like Sequence O21 280 PHEOOO2227 2 334 maize protease inhibitor like 2 sequence O22 281 PHEOOO2232 2 339 Soybean ABA-responsive element binding protein 2 (AREB2) like sequence US 2008/0148432 A1 Jun. 19, 2008

TABLE 1-continued

PEP NUC SEQ SEQ ID ID Base NO NO GENEID Vector PPROTEIN NAME O23 282 PHE0002233 2340 Arabidopsis ABA-responsive element binding protein 3 (AREB3) like sequence O24 283 PHEOOO2238 2345 soybean CPRD14 like 1 sequence O25 284 PHEOOO2239 2346 Brassica napus CPRD14 like 1 sequence O26 285 PHEOOO2240 2347 soybean CPRD14 like 2 sequence O27 286 PHEOOO2242 2349 maize CPRD14 like 1 sequence O28 287 PHEOOO2244. 2351 Arabidopsis drought inducible heat shock transcription factor like sequence O29 288 PHEOOO2246 2353 Soybean protein phosphatase 2C like sequence O3O 289 PHEOOO2254 2361 soybean CPRD12 like 3 sequence O31 290 PHE0002268. 2371 Soybean calcium-dependent protein kinase like 1 sequence O32 291 PHEOOO2269 2372 rice calcium-dependent protein kinase like 1 Sequence O33 292 PHEOOO2272 2375 rice calcium-dependent protein kinase like 4 Sequence O34 293 PHE0002298 2400 4 rice Dehydrin ERD10 like 1 sequence O35 294 PHEOOO2299 2401 rice drought-induced S-like ribonuclease like 1 sequence O36 295 PHEOOO2321 2422 maize Dil.9 like sequence O37 296 PHEOOO2331, 2432 soybean PIP like 1 sequence O38 297 PHEOOO234O 2441 soybean MIP4 like 1 sequence O39 298 PHEOOO2354 2455 maize glutathione transferase like 2 sequence 040 299 PHEOOO2357 2458 maize 8-oxoguanine-DNA glycosylase like Sequence O41 300 PHEOOO2361. 2462 rice CEO1 like sequence O42 301 PHEOOO2362 2463 maize dolichyl-phosphate beta glucosyltransferase like sequence O43 302 PHEOOO2364 2465 maize glutathione reductase (GR2) like 1 Sequence O44 303 PHEOOO2365. 2466 soybean glutathione reductase (GR2) like 1 Sequence O45 304 PHEOOO2367 2468 rice glutathione reductase (GR2) like 1 Sequence O46 305 PHEOOO2370 2471 Soybean glutathione-peroxidase like 1 Sequence O47 306 PHEOOO2372 2473 maize glutathione-peroxidase like 2 sequence O48 307 PHEOOO2373 2474 maize glutathione-peroxidase like 3 sequence O49 308 PHEOOO2383. 2484 maize T-complex polypeptide 1 alpha Subunit like sequence O50 309 PHEOOO2390 2491 Soybean methionine Sulfoxide reductase (mSr) like sequence O51 310 PHE0002414 2.514 Arabidopsis GAD2 O52 311 PHEOOO2431. 2531 Rice Leucine Zipper Protein similar to Atl03 and PNIL34 O53 312 PHEOOO2447 2547 soybean arabidopsis-heat-shock-TF like 1 Sequence O54 313 PHEOOO2449 2549 Soybean heat shock factor 6 like 1 sequence O55 314 PHEOOO2459 2559 rice heat shock TF like sequence O56 315 PHEOOO2461. 2561 soybean HSF1 like 1 sequence 057 316 PHEOOO2462 2562 soybean HSF4 like 1 sequence O58 317 PHEOOO2464. 2564 soybean HSF4 like 3 sequence O59 318 PHEOOO2470 2570 Soybean hsp17.4 like 2 sequence O60 319 PHEOOO2471. 2571 Soybean hsp17.4 like 3 sequence O61 320 PHE0002484 2584 Corn Sucrose Synthase O62 321 PHEOOO2486 2.586 Corn Putative protein phosphatase 2A regulatory Subunit B O63 322 PHEOOO2489 2589 Arabidopsis CycD2 O64 323 PHEOOO2490. 2590 Arabidopsis CycD3 O65 324 PHEOOO2490 3963 16 Arabidopsis CycD3 O66 325 PHEOOO2491. 2591 Arabidopsis CycB1 O67 326 PHEOOO2498 2.598 Corn Protein Kinase like protein O68 327 PHEOOO25O2 2602 Corn 20 KDA CHAPERONIN, CHLOROPLAST PRECURSOR (PROTEIN CPN21) O69 328 PHEOOO2506. 2606 Corn protein of unknown function wiPFAM domain similar to human Reticulon and may associate with ER O70 329 PHEOOO2507 2607 Corn Transfactor like protein O71 330 PHEOOO2514 2614 Corn branched-chain alpha keto-acid dehydrogenase E1 alpha subunit US 2008/0148432 A1 Jun. 19, 2008

TABLE 1-continued

PEP NUC SEQ SEQ ID ID Base NO NO GENEID Vector PPROTEIN NAME O72 331 PHEOOO2515 2615 Corn haloacid dehalogenase-like hydrolase Putative ripening related protein O73 332 PHEOOO2530 2630 Rice Dof zinc finger protein O74 333 PHEOOO2532 2632 Corn ribosomal protein L1p/L10e family O75 334 PHEOOO2536 2636 Rice auxin response transcription factor 3-like protein O76 335 PHEOOO2540. 2640 Oryza Saiva auxin response factor 2 077 336 PHEOOO2547. 2647 Corn homolog to Arabidopsis protein wi hydrolase fold O78 337 PHEOOO2551 2651 Corn TF with 3 leucine Zippers & PFAM mTERF domain O79 338 PHEOOO2552 2652 Corn protein with Duf210 PFAM domain O80 339 PHEOOO2565. 2664 Corn metallothionein-like protein O81 340 PHEOOO2581. 2680 maize hsp60 like 1 sequence O82 341 PHEOOO2582. 2681 maize hsp60 like 2 sequence O83 342 PHEOOO2583 2682 maize hsp60 like 3 sequence O84 343 PHEOOO2586. 2685 rice hsp60 like 1 sequence O85 344 PHEOOO2588. 2687 Corn protein with “Universal Stress Protein Family PFAM domain O86 345 PHEOOO2591. 2690 Corn protein similar to Yersinia pestis membrane protein and Bacilius Subtilis yuxK O87 346 PHEOOO2596 2695 Corn Myb related Transcription Factor O88 347 PHEOOO2604 2703 Rice putative putative photoreceptor interacting protein-like protein O89 348 PHEOOO2608. 2707 Arabidopsis sigma factor 2 O90 349 PHEOOO2615 2714 corn sigA binding protein 2 O91 350 PHEOOO2622 2739 Arabidopsis GLK1 O92 351 PHEOOO2629 2748 rice trehalose-6-P synthase like 1 sequence O93 352 PHEOOO2634 2753 rice DL-glycerol-3-phosphatase like 1 Sequence O94 353 PHEOOO2639 2758 rice GH3 protein 8 O95 354 PHEOOO2640 2759 rice GH3 protein 3 O96 355 PHEOOO2643. 2762 rice GH3 protein 2 O97 356 PHEOOO2644 2763 rice GH3 protein 9 O98 357 PHEOOO2649 2768 wheat AGL21-like 1 O99 358 PHEOOO2661. 2781 5 Synechocystis sp. PCC 6803 ADP-glucose pyrophosphorylase OO 359 PHEOOO2664. 2787 6 rice ADP-glucose pyrophosphorylase 5 O1 360 PHE0002688. 2821 rice beta-3 tubulin like 1 sequence O2 361 PHE0002689 2822 rice beta-3 tubulin like 2 sequence O3 362 PHEOOO2690 28.23 Corn protein similar to cell division related protein kinase 04 363 PHEOOO2703. 2836 rice VTC2 like 1 sequence 05 364 PHE0002710 2843 Zea Mays cytoplasmic malate dehydrogenase O6 365 PHEOOO2715 2848 Oryza sativa putative thiolase 07 366 PHE0002717 2850 Corn Translation Elongation factor EF1-beta O8 367 PHE0002721. 2854 4 Maize fructose-bisphosphate aldolase O9 368 PHE0002724. 2859 Corn L19 Like ribosomal protein 10 369 PHEOOO2728 2861 maize Sucrose transport protein SUC2 like 1 Sequence 11 370 PHEOOO2729 2863 Corn 60S ribosomal protein L10 (probable transcription factor) 12 371 PHEOOO2733 2866 Corn Ribosomal protein S11 13 372 PHEOOO2734 2867 Corn Ribosomal protein S12 14 373 PHEOOO2749 2882 rice Sucrase-like 1 sequence 15 374 PHEOOO2751. 2884 Corn Ribosomal protein S14 16 375 PHEOOO2752 2885 Corn Homolog to putative 40S Ribosomal protein 17 376 PHEO002771. 2904 Zea mays beta-glucosidase aggregating factor precursor 18 377 PHEOOO2790 2925 rice CYP72A5 like 1 sequence 19 378 PHEOOO2846. 2981 Zea MayStrehalose-6-phosphate phosphatase 20 379 PHEOOO2864. 2999 soy CDKA 8 21 380 PHEOOO2869 3004 corn CDKD 12 22 381 PHE0002875 3010 Corn homolog to Arabidopsis unknown expressed protein 23 382 PHEOOO2889 3024 soydsPTP3 24 383 PHEOOO2896 3031 rice disPTP1 25 384 PHEOOO2918. 3053 Oryza Saiiva putative Hexose Transporter IV (distant homology to Yeast Maltose permease) US 2008/0148432 A1 Jun. 19, 2008 16

TABLE 1-continued

NUC SEQ ID Base NO GENEID Vector PPROTEIN NAME 385 PHEOOO2946. 3096 Zea mays Putative Polyamine Transporter 386 PHEOOO2963. 31.13 Zea Mays Dual Specificity Phosphatase I (similar to human YVH1) 28 387 PHEOOO2966 3116 Oryza Saiiva Dual Specificity Phosphatase II 29 388 PHEOOO2984 3134 Zea mass 3-phosphoinositide-dependent protein kinase 30 389 PHEOOO3061. 3211 Oryza sativa Putative integral membrane protein 31 390 PHEOOO3074 3224 Zea Mays RNA Binding Protein 32 391 PHEOOO3101 3969 Soy nonsymbiotic hemoglobin 33 392 PHEOOO3101 76 Soy nonsymbiotic hemoglobin 34 393 PHEOOO3124 3270 soy adenylylsulfate kinase like 35 394 PHEOOO3138 3292 rice fructokinase 1 36 395 PHEOOO3139 3295 corn fructokinase 1 37 396 PHEOOO3190 3389 soy G1225-like 2 38 397 PHEOOO3196 3395 Arabidopsis seven-in-absentia 1 39 398 PHEOOO31.98 3397 corn seven-in-absentia 5 40 399 PHEOOO3211 3417 Arabidopsis AVP1 41 400 PHEOOO3211 3967 Arabidopsis AVP1 42 401 PHE0003217 3423 soy G200-like 13 43 402 PHEOOO3224 3432 corn-CGPG1897-like5 403 PHEOOO3228 3444 soy SAG13 full length 45 404 PHEOOO3229 3445 soy SAG13 truncated 46 405 PHEOOO3237 3453 rice-CGPG1264-like1 47 406 PHEOOO3240 3456 soy-CGPG1857-like1 48 407 PHEOOO3243 3459 corn-CGPG867-like4 49 408 PHEOOO3253 3470 soy-CGPG1276-like1 50 409 PHEOOO3257 3474 soy-CGPG1294-like2 51 410 PHEOOO3273 3492 soy-CGPG1287-like1 52 411 PHEOOO3276. 3495 Arabidopsis dehydrogenase 53 412 PHEOOO3282 35O1 corn-CGPG241-like3DUF6) S4 413 PHEOOO3286 3505 rice CTP synthase 55 414 PHEOOO3287 3506 corn CTP synthase 56 415 PHEOOO3304 3524 corn unknown protein 57 416 PHEOOO3309 3528 soy-CGPG1307-like1 58 417 PHEOOO3312. 3531 Arabidopsis nitrate transporter 59 418 PHEOOO3321 3540 Arabidopsis PDK1 like 2 60 419 PHEOOO3327 3546 corn NEK like-1 61 420 PHEOOO3328 3547 corn NEK like-2 62 421 PHEOOO3330 3549 Arabidopsis eIFiso4E like p28 subunit 63 422 PHEOOO3333 3552 soy eIF4E like 1 64 423 PHEOOO3333 4250 17 soy eIF4E like 1 65 424 PHEOOO3336 3555 Arabidopsis eIF4E 66 425 PHE0003353. 3572 corn seven-in-absentia 5 C57S 67 426 PHEOOO3361 3584 malate dehydrogenase 68 427 PHEOOO3362. 3585 hexokinase 69 428 PHEOOO3364. 3587 aminotransferase-like protein 70 429 PHE0003369. 3592 glyceraldehyde 3-phosphate dehydrogenase 71 430 PHEOOO3373 3596 putative purple acid phosphatase 72 431 PHEOOO3381 3604 hypothetical protein2 73 432 PHEOOO3385 3608 expressed protein2 74 433 PHEOOO3391 3614 serine/threonine protein kinase like protein 75 434 PHEOOO3392 3615 lipase-like protein 76 435 PHEOOO3393 3616 Putative Squalene monooxygenase 77 436 PHEOOOO132 222 FUS5 78 437 PHE0000285 375 Sorghum mLIP15 79 438 PHEOOOO668 771 Arabidopsis glycerol-3-phosphate acyltransferase-D00673 8O 439 PHE0000790 863 soyagl1 1-like 1 delta MADS-box 81 440 PHEOOO1074 1164 Arabidopsis NAC domain transcription factor 82 441 PHEOOO1078 11.68 Arabidopsis NAC domain transcription factor 83 442 PHEOOO1079 1169 Arabidopsis NAC domain transcription factor 84 443 PHEOOO1082 1172 Arabidopsis NAC domain transcription factor 85 444 PHEOOO1085. 1175 Arabidopsis myc domain transcription factor 86 445 PHEOOO1113 1203 Arabidopsis serine glyoxylate aminotransferase-AB048945 87 446 PHEOOO1136 1226 rice aspartate aminotransferase-D14673 88 447 PHEOOO1142 1232 corn caffeoyl-CoA 3-O-methyltransferase 89 448 PHEOOO1216 1306 maize G1836-like 1 90 449 PHEOOO1240. 1330 Inorganic pyrophosphatase 91 450 PHEOOO1243. 1333 Tubby Protein US 2008/0148432 A1 Jun. 19, 2008

TABLE 1-continued

PEP NUC SEQ SEQ ID ID Base NO NO GENEID Vector PPROTEIN NAME 92 451 PHEOOO1248. 1339 Arabidopsis hypothetical protein 93 452 PHEOOO1257 1348 Arabidopsis NADH:ubiquinone oxidoreductase 94 453 PHEOOO1263. 1354 Arabidopsis hypothetical protein NM 114619) 95 454 PHEOOO1266 1357 corn hypothetical protein 96 455 PHEOOO1283 1373 Arabidopsis Asp1 97 456 PHEOOO1286 1376 Arabidopsis putative inositol hexaphosphate kinase 98 457 PHEOOO1301. 1391 Arabidopsis Pra2-like protein-AAF97325 99 458 PHEOOO1304 1394 corn Pra2-like protein 1 200 459 PHEOOO1316. 1406 rice CGPG 1145-like protein 1 201 460 PHEOOO1317 1407 rice CGPG 1145-like protein 2 202 461 PHEOOO1319. 1409 Arabidopsis hypothetical protein 203 462 PHEOOO1323. 1413 Arabidopsis hypothetical protein NM 111447) 204 463 PHEOOO1324 1414 Arabidopsis expressed protein 205 464 PHEOOO1466 1558 Arabidopsis thaliana ROOT HAIRLESS 1 (RHL1)-AAC23500.1 206 465 PHEOOO1564. 1663 4 Xanthomonas campestris asparagine synthase 207 466 PHEOOO2094 2.204 Corn serine/threonine kinase 208 467 PHEOOO2121 2229 corn G664-like 4 209 468 PHEOOO2225 2332 rice drought-induced protein Di19-like Sequence 210 469 PHEOOO2226. 2333 maize protease inhibitor like 1 sequence 211 470 PHEOOO2293 2395 Soybean drought-inducible cysteine proteinase like 1 sequence 212 471 PHEOOO2338 2439 maize MIP3 like 3 sequence 213 472 PHEOOO2524. 2624 Rice putative thioredoxin like protein 214 473 PHEOOO2567 2666 Arabidopsis VPE1 215 474 PHEOOO2607 2706 Arabidopsis Sigma factor 1 216 475 PHEOOO2609 2708 Arabidopsis sigma factor 3 217 476 PHEOOO2625 2744 rice waxy like 1 sequence 218 477 PHEOOO2939 3O89 Oryza Saiiva Proline oxidase (proline dehydrogenase) 219 478 PHEOOO3057. 3207 Schizosaccharomyces pombe orb6 220 479 PHEOOO3182 3341 Neurospora crassa Glycogen synthase like Sequence 221 480 PHEOOO3232 3448 soy-CGPG1264-like3 222 481 PHEOOO3234 3450 rice-CGPG1464-like3 223 482 PHEOOO3239 3455 corn-CGPG1857-like2 224 483 PHEOOO3246 3462 Arabidopsis hypothetical protein 225 484 PHEOOO3259 3476 Arabidopsis Microtubule associated protein 226 485 PHEOOO3267 3486 Arabidopsis Glycine-tRNA 227 486 PHEOOO3269 3488 Arabidopsis unknown protein 228 487 PHEOOO3274 3493 Arabidopsis Unknown protein 229 488 PHEOOO3285 3504 rice-CGPG566-like1 230 489 PHEOOO3289 3508 8 corn aluminium-induced protein 231 490 PHEOOO3311 3530 Arabidopsis hypothetical protein 232 491 PHEOOO336O 3.583 3-ketoacyl-ACP synthase 233 492 PHEOOO3365 3588 putative chlorophyll synthase 234 493 PHEOOO3374 3597 putative xyloglucan endotransglycosylase 235 494 PHEOOO3375 3598 Tryptophan synthase alpha chain 236 495 PHE0003378. 36O1 MN3 237 496 PHEOOO3379 3602 hypothetical protein1 238 497 PHE0003383 3606 expressed protein1 239 498 PHEOOO3387 3610 4 nodulin-like protein 240 499 PHEOOO3401 3624 corn Sterol 5 alpha desaturase 241 500 PHEOOO3417 3657 19 soy G1634-like 1 242 501 PHEOOO3418 3658 19 Arabidopsis isocitrate 243 502 PHEOOO3433 3642 corn dehalogenase-phosphatase 1 244 503 PHEOOO3450 3681 soy G1274 like 1 245 504 PHEOOO3458. 3689 corn G1660 like 1 246 505 PHEOOO3459 3690 corn G1730 like 1 247 506 PHEOOO3476 3707 soy G1988 like 1 248 507 PHEOOO3484 3730 corn G2035 like1 249 508 PHEOOO3491 3737 soy G2063 like2 250 509 PHEOOO3499 3745 rice G2207 like2 251 510 PHE0003518 3764 soy G2505 like1 252 511 PHEOOO3524 3770 soy G2536 like1 253 512 PHEOOO357O 3816 soy G922 like1 US 2008/0148432 A1 Jun. 19, 2008 18

TABLE 1-continued

PEP NUC SEQ SEQ ID ID Base NO NO GENEID Vector PPROTEIN NAME 254 513 PHEOOO3576 3822 1 soy G975 like1 255 514 PHEOOO3583 3829 1 corn G728 like1 256 515 PHEOOO3587 3833 1 soy G3083 like1 257 516 PHEOOO3599 3845 1 corn G2981 like1 258 517 PHEOOO3600 3846 4 corn R-S 259 518 PHEOOO3630 3888 1 corn G1206 like 2 260 519 PHEOOO3641. 3899 4 corn G2998 like2 261 520 PHEOOO3644 3902 1 corn G303 like2 262 521 PHEOOO3650 3908 1 soy G355 like2 263 522 PHEOOO3678 3936 1 corn G1052 like2 264 523 PHEOOO3686 3961 16 soy NDPK2 265 524 PHEOOO3740 4042 16 soy G481-like 3 266 525 PHEOOO3742. 4044 4 corn UDP-glucose 4-epimerase CGPG1003 like2 267 526 PHEOOO3743. 4046 4 rice UDP-glucose 4-epimerase CGPG1003 like1 268 527 PHE0003825. 4.192 4 soy G1412 like1 269 528 PHEOOO3839 4208 4 rice G2604 like1 270 529 PHE0003885 4266 4 wheat hemoglobin 271 530 PHE0003886 4267 4 Arabidopsis hemoglobin-like 1 272 531 PHEOOO3927 4321 4 Zea mays AIH-like 273 532 PHEOOO3959 4544 4 Glycine max CGPG7857 pseudo-response regulator 274 533 PHEOOO3961. 4662 14 maize Kas I 275 534 PHEOOO3965. 4553 4 corn ABI5 like 276 535 PHEOOO3966 4554 4 Corn AtlPS1-like 277 536 PHEOOO3977 4565 4 corn AfMONFEEDOO1213 uroporphyrinogen III synthase 278 537 PHEOOO3978. 4566 4 corn AfvONFEEDOOO902 proteophosphoglycan 279 538 PHEOOO3982 4569 4 corn AfMONFEED000317 Serine carboxypeptidase II-1 280 539 PHEOOO3993 4579 4 corn AfMONFEEDOO1219 putative Ca2+H---exchanging protein 281 540 PHEOOO3994 4580 4 corn AfMONFEED001169 putative pathogenesis related protein 282 541 PHEOOO3996 4582 4 corn AfMONFEEDOOO906 auxin-induced protein-related 283 542 PHEOOO3997 4583 4 corn AfMONFEEDOO1366 isoflavone reductase homolog 284 543 PHEOOO3999 4585 4 corn AfMONFEEDOOO689 proline-rich protein 285 544 PHEO004OOO 4586 4 corn AfMONFEED000920 nodulin-related protein 286 545 PHEOOO4002 4588 4 corn AfMONFEEDOO1475 hypothetical protein 287 546 PHEO004003 4589 4 corn AfMONFEED000648 acyltransferase 288 547 PHEO004004 4590 4 corn AfMONFEED000503 pathogenesis related protein 4 289 548 PHEO004005 4591 4 corn AfMONFEEDOO1364 unknown protein 290 549 PHEO004006 4592 4 corn AfMONFEED000623 hydrolase 291 550 PHEO004009 4595 4 corn AfMONFEED000074 unknown protein 292 551 PHEOOO4010 4596 4 corn AfMONFEED000152 Myb-like DNA binding domain 293 552 PHEO004011 4597 4 corn AfMONFEED001001 ACT domain containing protein 294 553 PHEOOOO788 861 4 corn agl1 1-like 1 delta C-terminus 295 554 PHEOOOO789 862 4 corn agl1 1-like 1 delta K-box 296 555 PHE0000795 868 1 rice MADS3 delta K-box-L37528 297 556 PHEOOOO881 964 1 Synechocystis sp. PCC 6803 Hik33 298 557 PHEOOO1226 1316 1 wheat catalase-like 1 299 558 PHEOOO1253. 1344 1 soy arabinoglactan-protein 1 300 559 PHEOOO1335. 1426 1 rice SNF1-like protein 8 OsPK7 BAA83689 301 560 PHEOOO1381. 1473 1 rice ANR1-like 302 561 PHEOOO1521. 1612 1 soy LPE1-like permease 2 303 562 PHEOOO1630. 1741 1 maize low temperature and Salt responsive protein LTI6A-like 2 sequence 304 563 PHEOOO1659 1770 1 corn SDD1-like 1 pre-pro domain 305 564 PHEOOO2O79. 21.89 1 rice seco1 306 565 PHEOOO2185 2292 1 rice Hsp17.7 like 1 sequence US 2008/0148432 A1 Jun. 19, 2008

TABLE 1-continued

PEP NUC SEQ SEQ ID ID Base NO NO GENEID Vector PPROTEIN NAME 307 566 PHEOOO2229 2336 rice protease inhibitor like 1 sequence 308 567 PHEOOO2392. 2493 rice copper chaperone like 1 sequence 309 568 PHEOOO2410 2510 E. CoigadA 310 569 PHE0002411 2511 Synechocystis sp. PCC 6803 gad 311 570 PHEOOO2485 2.585 4 Corn Amino Acid Transport Protein 312 571 PHEOOO2496. 2596 Corn Glutamate-1-semialdehyde aminotransferase 313 572 PHE0002611 2710 soy sigma factor 1 314 573 PHE0002616. 2723 Arabidopsis proline:glycine betaine transporter 1 315 574 PHEOOO2670 2793 5 Streptomyces coelicolor glucose-1-phosphate adenylyltransferase 316 575 PHE0002727 2860 rice Sucrose synthase-1 like 2 sequence 317 576 PHEOOO295O 3100 Yeast protein involved in resistance to H2O2 318 577 PHEOOO2997 3147 Yeast Protein of unknown function, has WD (WD-40) repeats 319 578 PHEOOO3O11 3161 Yeast Protein (wi Carbonic anhydrase PFAM domain) involved in protection against oxidative damage 320 579 PHEOOO3O21, 3171 Yeast protein of unknown function (may be involved in cell damage) 321 580 PHEOOO3200 3399 Arabidopsis seven-in-absentia 2 322 581 PHEOOO3227 .3443 16 soy G1820 like 323 582 PHEOOO3261 3478 rice-CGPG1517-like2 324 583 PHEOOO3270 3489 soy-CGPG 1191-like1 325 584 PHEOOO3277 3496 soy-CGPG1430-like1 326 585 PHEOOO3352 3571 Corn ethylene receptor 1 327 586 PHEOOO3366 3589 transaldolase 328 587 PHEOOO3370 3593 4 Circulin B 329 588 PHEOOO3526 3772 soy G2567 like2 330 589 PHEOOO3581 3827 rice G2603 like2 331 590 PHEOOO3681 3942 4 rice G1792-like 3 332 591 PHEOOO3683 3944 16 soy SUC1-like 1 333 592 PHEOOO3683 3945 18 soy SUC1-like 1 334 593 PHEOOO3815 4288 4 Synechocystis NblS-like 335 594 PHEOOO3986. 4573 4 corn AfMONFEEDOO1223 putative stearoyl acyl-carrier protein desaturase 336 595 PHEOOO3989 4575 4 corn AfMONFEEDO00878 remorin-like protein 337 596 PHEOOO3990 4576 4 corn AfMONFEEDOO1231 unknown protein 338 597 PHEOOO3995 4581 4 corn AfMONFEEDOO 1033 putative succinate dehydrogenase 339 598 PHEO004022 4657 4 Arabidopsis Alfin-like 340 599 PHEOOOO667 770 chimeric glycerol-3-phosphate acyltransferase 341 600 PHEOOOO784 857 Arabidopsis ag|11 delta MADS-box 342 601 PHEOOOO880 963 Nostoc sp. PCC7120 Hik33 343 602 PHEOOOO883 966 Synechocystis sp. 6803 Rerl 344 603 PHEOOO1063. 1153 yeast SNF4-Z72637 345 604 PHEOOO1065. 1155 yeast SIP2-Z72730 346 605 PHEOOO1069 1159 Arabidopsis bZIP domain transcription factor 347 606 PHEOOO1076 1166 Arabidopsis NAC domain transcription factor 348 607 PHEOOO1080 1170 Arabidopsis AP2 domain transcription factor 349 608 PHEOOO1087 1177 Arabidopsis homeodomain transcription factor 350 609 PHEOOO1094. 1184 Arabidopsis zinc finger protein 351 610 PHEOOO1100 1190 Arabidopsis bZIP domain transcription factor 352 611 PHEOOO1112 1202 yeastalanine aminotransferase 2-CAA88665 353 612 PHEOOO1131. 1221 5 yeast acetohydroxyacid reductoisomerase AAB677S3 354 613 PHEOOO1143 1233 corn putative isoprenylated protein 355 614 PHEOOO1149 1239 corn HvB12D homolog 356 615 PHEOOO1169 1259 Arabidopsis TFL2-AF387639 357 616 PHEOOO1176 1266 5 Synechocystis sp. PCC 6803 heme oxygenase-1651897 358 617 PHEOOO1177 1267 5 Nostoc sp. PCC 7120 heme oxygenase 17132210 359 618 PHEOOO1184 1274 1 rice SHY2-like 1-BAA92982 360 619 PHEOOO1256 1347 1 Arabidopsis protease HhoA precursor 361. 620 PHEOOO1264 1355 1 Arabidopsis hypothetical protein 362 621 PHEOOO1265 1356 1 Arabidopsis hypothetical protein NM 105159) US 2008/0148432 A1 Jun. 19, 2008

TABLE 1-continued

PEP NUC SEQ SEQ ID ID Base NO NO GENEID Vector PPROTEIN NAME 363 622 PHEOOO1268- 1359 Arabidopsis hypothetical protein 364 623 PHEOOO1270. 1361 yeasterol-CAA90553 365 624 PHEOOO1311 1401 Arabidopsis expressed protein NM 126228 366 625 PHEOOO1322. 1412 Arabidopsis expressed protein 367 626 PHEOOO1519 1610 rice WD domain protein 368 627 PHEOOO1523. 1614 rice LPE1-like permase 1-BAB61205 369 628 PHEOOO1527. 1618 Nostoc sp. PCC 7120 GDH 37O 629 PHEOOO1528 1619 Nostoc punctiforme GDH 371 630 PHEOOO1533 1624 Streptomyces coelicolor GDH 372 631 PHEOOO1541 1632 rice glutamate dehydrogenase-15787849 373 632 PHEOOO1567. 1666 Bacilitis halodurans asparagine synthase 10174030 374 633 PHEOOO1568 1667 4 Corynebacterium glutamictim Asparagine synthase-19551250 375 634 PHEOOO1611 1722 rice translation initiation factor 3 delta subunit like sequence 376 635 PHEOOO1632 1743 arabidopsis magnesium/proton exchanger AtMHX 377 636 PHE0002029 2140 Corn Axi 1 378 637 PHEOOO2O78 2188 yeast Sec61 379 638 PHEOOO2O84 2.194 Corn cyclic nucleotide and calmodulin regulated ion channel 380 639 PHEOOO2089. 2199 Agrobacterium timefaciens hypothetical protein 381 640 PHEOOO2090 2200 Nostoc sp. PCC 7120 stress induced hydrophobic peptide 382 641 PHE0002151 2259 Corn Kaurene Synthase 383 642 PHEOOO21942301 maize Betaine-aldehyde dehydrogenase like 1 Sequence 384 643 PHEOOO2483 2583 maize heat shock TF like sequence 385 644 PHEOOO2494 2594 Arabidopsis ascorbate oxidase 386 645 PHEOOO2509 2609 cORN Hsp20/alpha crystallin family 387 646 PHEOOO2523 2623 Corn putative thioredoxin protein 388 647 PHEOOO2527 2627 Hypothetical Nostoc protein similar to cobW 389 648 PHEOOO2546 2646 Corn dual-specificity protein phosphatase-like protein 390 649 PHEOOO2557 2657 soy G1792-like 10 391 650 PHEOOO2566 2665 Ribosomal protein L39 392 651 PHE0002614 2713 Brassica naptis sigA binding protein 2 393 652 PHEOOO2648. 2767 soy AGL21-like 2 394 653 PHEOOO2652. 2771 Corn GDP-mannose pyrophosphorylase A 395 654 PHEOOO2791. 2926 rice CYP72A5 like 2 sequence 396 655 PHE0002849 2984 Oryza Saiiva Putative Glucosyl Transferase 397 656 PHE0002855 2990 Synechocystis sp unknown protein wit ABC1 PFAM domain (putative novel chaperonin) 398 657 PHE0002856. 2991 Nostoc sp unknown protein wit ABC1 PFAM domain(putative novel chaperonin) 399 658 PHEOOO2886 3021 Brassica dsPTP1 400 659 PHEOOO2888. 3023 soydsPTP2 401 660 PHEOOO2893 3028 Corn dSPTP3 402 661 PHEOOO2901 3.036 Yeast GAT2 GATA Zinc Finger TF 403 662 PHEOOO2906. 3041 Zea Mays Zinc Finger Transcription Factor IV 404 663 PHEOOO2910 3045 Yeast IKS1 405 664 PHEOOO2923 3058 Yeast GGA1 protein (mediator of protein traficking between Trans golgi network and vacoule) 406 665 PHEOOO2928 3065 Yeast Zinc Finger protein (DNA damage responsive repressor of PHR1) 407 666 PHEOOO2941. 3091 Yeast SET6p putative Transcription Factor 408 667 PHEOOO2947 3097 Zea Mays Sugar & other (polyamine) transporter like protein 409 668 PHEOOO2948 3098 Yeast Xylulokinase 410 669 PHEOOO2987 3137 Yeast oxidoreductase of unknown function (PFAM NAD-binding Rossmann fold & C terminal alphabeta domain) 411. 670 PHEOOO3017 3167 Yeast Transketolase 2 412 671 PHEOOO3O18, 31.68 Yeast Ubiquitin polyprotein 413 672 PHEOOO3O23 3173 Yeast unknown protein (uncharacterized protein family) US 2008/0148432 A1 Jun. 19, 2008

TABLE 1-continued

PEP NUC SEQ SEQ ID ID Base NO NO GENEID Vector PPROTEIN NAME 414 673 PHEOOO3O25 31 75 Yeast Protein of unknown function, (similar to mouse MPV17 a putative integral membrane peroxisomal protein) 415 674 PHEOOO3026 3176 Yeast Protein of unknown function, putative paralog of Ecm4p, a cell wall biogenesis protein 416 675 PHEOOO3027 .3177 Yeast Protein of unknown function (Pfam Domain YeF-related protein N-terminus) 417 676 PHEOOO3O28 3178 Yeast Potential alpha-ketoisocaproate reductase 418 677 PHEOOO3O3O 318O Yeast Yapsin 6, GPI-anchored aspartyl protease 419 678 PHEOOO3062 3212 soy G571 42O 679 PHEOOO3O88 3240 Arabidopsis AtPK1 421 680 PHEOOO3O89 3237 Arabidopsis AtPK19b 422 681 PHEOOO3090 3238 soy S6K1 423 682 PHEOOO3091 3239 corn S6K1 424 683 PHEOOO3102 3244 6 Clostridium acetobutyllicum fructokinase 425 684 PHEOOO3102 3579 16 Clostridium acetobutyllicum fructokinase 426 685 PHEOOO3121. 3267 Arabidopsis cyt P450 like 427 686 PHEOOO3130 3276 Arabidopsis glucosyltransferase-like 428 687 PHEOOO3134 3280 Arabidopsis microtubule-associated protein EB1-like protein like 429 688 PHEO003194 3393 corn seven-in-absentia 2 430 689 PHEOOO3195 3394 soy seven-in-absentia 1 431 690 PHEOOO3199 3398 rice seven-in-absentia 2 432 691 PHEOOO32O1 3400 soy seven-in-absentia 4 433 692 PHEOOO3208 3410 7 soy triose phosphate translocator 1 434 693 PHEOOO3213 3419 yeast TAT2 435 694 PHEOOO3214 3420 yeast CUP1a 436 695 PHEOOO3222 3428 Corynebacterium glutamicum glutamate 5 kinase 437 696 PHEOOO3238 3454 Arabidopsis fructose-bisphosphate aldolase 438 697 PHEOOO3242 3458 Arabidopsis ABC transporter 439 698 PHEOOO3251 3468 Arabidopsis unknown protein 440 699 PHEOOO326O 3477 soy-CGPG 1517-like 441 700 PHEOOO3272 3491 Arabidopsis hypothetical protein 442 701 PHEOOO3275 3494 Brassica-CGPG1391-like1 443 702 PHEOOO3302 3523 Arabidopsis hypothetical protein 444 703 PHEOOO33O8 3527 Arabidopsis Yippee putative zinc-binding protein 445 704 PHEOOO3315 3534 Arabidopsis unknown protein 446 705 PHEOOO3320 3539 Arabidopsis PDK1 like 447 706 PHEOOO3344 3562 corn ADC like-1 448 707 PHEOOO3347 3565 Arabidopsis GCN5 449 708 PHEOOO3354 3573 corn G571 long splice form A196P. A197P 450 709 PHEOOO3380 3603 putative laccase 451. 710 PHEOOO3388 3611 4 Caffeic acid 3-O-methyltransferase 452 711 PHEOOO3395 3618 4 hypothetical protein4 453 712 PHEOOO3397 3620 hypothetical proteinS 454 713 PHEOOO3403 3626 16 potato twin LOV protein 455 714 PHEOOO3411. 3634 Putative NAM protein 456 715 PHEOOO3414 3654 19 Streptococci is militansgtfA (dicot codon modified) 457 716 PHEOOO3415 3655 18 Synechococcus sp. PCC 7942 IctB with RuBisCO Small Subunit 1b CTP 458 717 PHEOOO3431. 3639 4 Streptomyces coelicolor trehalose synthase 1 459 718 PHEOOO3434 3643 1 corn dehalogenase-phosphatase 2 460 719 PHEOOO3436. 3645 1 Nostoc sp. PCC 7120 dehalogenase phosphatase 461 720 PHEOOO3437 3646 1 yeast dehalogenase-phosphatase 462 721 PHEOOO3447 3678 1 soy DUF296 463 722 PHEOOO3483 3729 1 soy G2035 like1 464 723 PHEOOO3519 3765 1 corn G2505 like2 465 724 PHEOOO3582 3828 1 soy G2999 like1 466 725 PHEOOO3586 3832 1 Brassica G2763 like1 467 726 PHEOOO3588 3834 1 soy G2776 like1 468 727 PHEOOO3598. 3844 1 soy G2981 like1 469 728 PHEOOO3606 3852 1 soy G.2898 like1 470 729 PHEOOO3613 3860 1 corn GAD1-1 471 730 PHEOOO3617 3866 1 Brassica G2839 like1 US 2008/0148432 A1 Jun. 19, 2008 22

TABLE 1-continued

PEP NUC SEQ SEQ ID ID Base NO NO GENEID Vector PPROTEIN NAME 1472 731 PHEOOO3632. 3890 1 rice G2982 like 1 1473 732 PHEO003636. 3894 1 soy G2992 like1 1474 733 PHEOOO3676. 3934 1 soy G1052 like2 1475 734 PHEOOO3682. 3943 4 wheat wipkA 1476 735 PHEOOO3788 4122 4 corn NAC1-like 1 1477 736 PHEOOO3899 4284 17 CGPG2101 Arabidopsis PHD-finger protein 1478 737 PHEOOO3948 4528 17 Arabidopsis CGPG3676 1479 738 PHEOOO3981. 4568 4 corn AfMONFEED000388 pyruvate kinase 1480 739 PHEOOO3984 4571 4 corn AfMONFEEDOOO668 unknown protein 1481 740 PHEOOO3998. 4584 4 corn AfMONFEED000499 putative indole-3- acetic acid-regulated protein 1482 741 PHEOOO40O8 4594 4 corn AtMONFEED000474 serine protease like protein

Selection Methods for Transgenic Plants with Enhanced on the ear, number of rows of kernels on the ear, kernel Agronomic Trait abortion, kernel weight, kernel size, kernel density and physi 0062. Within a population of transgenic plants regener cal grain quality. Although the plant cells and methods of this ated from plant cells transformed with the recombinant DNA invention can be applied to any plant cell, plant, seed or many plants that Survive to fertile transgenic plants that pro pollen, e.g. any fruit, vegetable, grass, tree or ornamental duce seeds and progeny plants will not exhibit an enhanced plant, the various aspects of the invention are preferably agronomic trait. Selection from the population is necessary to applied to corn, soybean, cotton, canola, alfalfa, wheat and identify one or more transgenic plant cells that can provide rice plants. In many cases the invention is applied to corn plants with the enhanced trait. Transgenic plants having plants that are inherently resistant to disease from the Mal de enhanced traits are selected from populations of plants regen Rio Cuarto virus or the Puccina sorghi fungus or both. erated or derived from plant cells transformed as described 0063. The following examples are included to demon herein by evaluating the plants in a variety of assays to detect strate aspects of the invention, those of skill in the art should, an enhanced trait, e.g. enhanced water use efficiency, in light of the present disclosure, appreciate that many enhanced cold tolerance, increased yield, enhanced nitrogen changes can be made in the specific aspects which are dis use efficiency, enhanced seed protein and enhanced seed oil. closed and still obtain a like or similar results without depart These assays also may take many forms including, but not ing from the spirit and scope of the invention. limited to, direct Screening for the trait in agreenhouse or field trial or by Screening for a Surrogate trait. Such analyses can be EXAMPLE 1. directed to detecting changes in the chemical composition, biomass, physiological properties, morphology of the plant. Plant Expression Constructs Changes in chemical compositions such as nutritional com position of grain can be detected by analysis of the seed A. Plant Expression Constructs for Corn Transformation composition and content of protein, free amino acids, oil, free fatty acids, starch or tocopherols. Changes in biomass char 0064. This example illustrates the construction of plas acteristics can be made on greenhouse or field grown plants mids for transferring recombinant DNA into plant cells which and can include plant height, stem diameter, root and shoot can be regenerated into transgenic plants of this invention. dry weights; and, for corn plants, ear length and diameter. 0065. A base plant transformation vectorpMON65154, as Changes in physiological properties can be identified by set forth in SEQ ID NO: 52768 was fabricated for use in evaluating responses to stress conditions, for example assays preparing recombinant DNA for transformation into corn using imposed stress conditions such as water deficit, nitro tissue using GATEWAYTM Destination plant expression vec gen deficiency, cold growing conditions, pathogen or insect tor systems (available from Invitrogen Life Technologies, attack or light deficiency, or increased plant density. Changes Carlsbad, Calif.). With reference to the elements described in in morphology can be measured by visual observation of Table 3 below, pMON65154 comprises a selectable marker tendency of a transformed plant with an enhanced agronomic expression cassette and a template recombinant DNA expres trait to also appear to be a normal plant as compared to sion cassette. The marker expression cassette comprises a changes toward bushy, taller, thicker, narrower leaves, striped CaMV 35S promoter operably linked to a gene encoding leaves, knotted trait, chlorosis, albino, anthocyanin produc neomycin phosphotransferase II (nptII) followed by a 3 tion, or altered tassels, ears or roots. Other selection proper region of an Agrobacterium tumefaciens nopaline synthase ties include days to pollen shed, days to silking, leaf extension gene (nos). The template recombinant DNA expression cas rate, chlorophyll content, leaf temperature, stand, seedling sette is positioned tail to tail with the marker expression vigor, internode length, plant height, leaf number, leaf area, cassette. The template recombinant DNA expression cassette tillering, brace roots, stay green, Stalk lodging, root lodging, comprises 5' regulatory DNA including a rice actin 1 pro plant health, barreness/prolificacy, green Snap, and pest resis moter, exon and intron, followed by a GATEWAYTM insertion tance. In addition, phenotypic characteristics of harvested site for recombinant DNA, followed by a 3' region of a potato grain may be evaluated, including number of kernels per row proteinase inhibitor II (pinII) gene. Once recombinant DNA US 2008/0148432 A1 Jun. 19, 2008 23 has been inserted into the insertion site, the plasmid is useful for plant transformation, e.g. by microprojectile bombard ment.

TABLE 3

FUNCTION ELEMENT REFERENCE Plant gene of interest Rice actin 1 promoter U.S. Pat. No. 5,641,876 expression cassette Rice actin 1 exon 1, intron 1 U.S. Pat. No. 5,641,876 enhancer Gene of interest AttR1 GATEWAYTM Cloning Technology insertion site instruction Manual CmR gene GATEWAYTM Cloning Technology instruction Manual cccdA, ccdB genes GATEWAYTM Cloning Technology instruction Manual attR2 GATEWAYTM Cloning Technology instruction Manual Plant gene of interest Potato pinII 3' region An et al. (1989) Plant Cell 1: 115-122 expression cassette Plant selectable CaMV 35S promoter U.S. Pat. No. 5,858,742 marker expression nptII selectable marker U.S. Pat. No. 5,858,742 caSSette nos 3' region U.S. Pat. No. 5,858,742 Maintenance in E. coli ColE1 origin of replication F1 origin of replication Blaampicillin resistance

0066. A similar base vector plasmid pMON72472 (SEQ ID NO: 52769) was constructed for use in Agrobacterium- TABLE 4 mediated methods of plant transformation similar to Base Vector ID pMON65154 except (a) the 5' regulatory DNA in the template Base Vector recombinant DNA expression cassette was a rice actin pro- or Corn moter and a rice actin intron, (b) left and right T-DNA border 1 MONf2472 sequences from Agrobacterium are added with the right bor- 2 MON65154 der sequence is located 5' to the rice actin 1 promoter and the 43 MON84109MON82O60 left border sequence is located 3' to the 35S promoter and (c) RN DNA is added to facilitate replication of the plasmid in both 7 MON81244 E. coli and Agrobacterium tumefaciens. The DNA added to MNI. the plasmid outside of the T-DNA border sequences includes 10 MON92667 an oriV- r wide host range origin of DNA replication functional 1211 MONf4582MON841.08 in Agrobacterium, a pBR322 origin of replication functional 13 MONf4579 in E. coli, and a spectinomycin/streptomycin resistance gene 14 bMONT4577Base Vector for selection in both E. coli and Agrobacterium. or Soybean 0067. Other base vectors similar to those described above 15 MONf45S2 were also constructed as listed in Table 4. See Table 4 for a 16 MONf4532 17 MON82O53 summary of base vector plasmids and base vector ID's which 18 MONf4537 are referenced in Table 1. Also see Table 5 for a summary of 19 MONf4536 regulatory elements used in the gene expression cassette for 2O MONf4548 these base vectors and SEQID NOs for elements.

TABLE 5

SEQ SEQ SEQ transit SEQ SEQ Vector promoter ID NO leader ID NO intron ID NO peptide ID NO terminator ID NO

pMON65154 P-Os. Act1-1:4:27 52789 L-Os. Act1-1:1:3 52777 I-OS.Act1-1:1:40 52.772 NONE T-St.Pisa-1:4:3 S28O1 pMON72472 P-Os. Act1-1:1:8 52788 L-OS.Act1-1:1:5 52.778 I-OS.Act1-1:1:3 52.771 NONE T-St.Pisa-1:4:1 S2800 pMONT4577 P-Hv.Per1-1:1:7 S2787 L-Hw.Per1-1:1:1 52776 I-Zm. DnaK-1:1:1 52773 NONE T-St.Pisa-1:4:1 S2800 pMONT4579 P-Zm.NAS2- 52793 L-Zm.NAS2- 52781 I-Zm. DnaK-1:1:1 52773 NONE T-St.Pisa-1:4:1 S2800 1:1:1 1:1:1 US 2008/0148432 A1 Jun. 19, 2008 24

TABLE 5-continued SEQ SEQ SEQ transi SEQ SEQ Vector broiloter ID NO leader ID NO intron ID NO peptide ID NO terminator ID NO MONT4582 P-ZPPDK- S2795 L-ZPPDK- S2782 I-Z.DnaK-1:1:1 S2773 NONE f T-St.PiSA-1:4:1 S2800 :1:12 :1:2 MONF6274 P-OS.GT1-1:1:10 S2790 L-OS.GT1-1:1:3 S2779 I-Z.DnaK-1:1:1 S2773 NONE f T-St.PiSA-1:4:1 S2800 MON81244 P-ZPPDK- S2794 L-ZPPDK- S2782 I-Z.DnaK-1:1:1 S2773 NONE f T-St.PiSA-1:4:3 52801 :1:10 :1:2 MON82O6O P-OS.Act1-1:1:8 52788 L-OS.Act1-1:1:5 52778 I-OS.Act1-1:1:3 52.771 NONE f T-St.PiSA-1:4:1 S2800 MON84109 P-OS.Act1-1:1:8 52788 L-OS.Act1-1:1:5 52778 I-OS.Act1-1:1:3 S2771 TS-Alt.ShkC- 52799 T-St.PS4-1:4:1 S2800 CTP2-1:1:1 MONf4430 P-OS.Act1-1:1:8 52788 L-OS.Act1-1:1:5 52778 I-OS.Act1-1:1:3 S2771 TS-Alt.ShkC- 52799 T-St.PS4-1:4:1 S2800 CTP2-1:1:1 MON84107 P-OS.GT1-1:1:18 52791 NONE I-Zm.DnaK-1:1:1. 52773 NONE f T-St.PiSA-1:4:1 S2800 MONT4575 P-Z.FDA-1:1:S 52792 L-Zm.FDA-1:1:1 S2780 I-Z.DnaK-1:1:1 S2773 NONE f T-St.PiSA-1:4:1 S2800 bMON92667 P-Zm.SZeinC1- 52797 L-Zm.SceinC1- S2783 I-Z.DnaK-1:1:1 S2773 NONE f T-St.PiSA-1:4:1 S2800 :: :1:1 MONT4532 P-CaMV.35S- 52786 NONE f NONE f NONE T-GbE6- 52798 ell- 3b:1:1 :1:11 MONT4552 P-CaMV.35S- 52786 NONE f NONE f TS-Alt.ShkC- 52799 T-Gb.E6- 52798 ell- CTP2-1:1:1 3b:1:1 :1:11 MON82OS3 P-CaMV.35S- 52786 NONE f NONE f NONE 52798 ell :1:11 MONT4537 P-At-RbCS4-1:1:2 52784 L-At-RbCS4-1:1:3 S2774 NONE f NONE 52798 pMON74536 P-BrSnap2-1:1:1 52785 L-Br. Snap2-1:1:1 52.775. NONE f NONE 52798 pMON74548 P-Gm. Sphas1- 52796 L-Gm. Sphas1- S2802 NONE f NONE 52798 :: 1:1:1 MONT4539 P-At-RbCS4-1:1:2 52784 L-At-RbCS4-1:1:3 S2774 NONE f TS-Alt.ShkC- 52798 CTP2-1:1:1 3b:1:1

0068 Plasmids for use in transformation of soybean were also prepared. Elements of an exemplary common expression vector plasmid pMON74532 are shown in Table 6 below.

TABLE 6

Function Element Reference Agro transformation B-ARGtu.right border Depicker, A. etal (1982) Mol Appl Genet 1:561-573 Antibiotic resistance CR-Ec.aadA-SPCSTR Repressor of primers from the ColE1 CR-Ec.rop plasmid Origin of replication OR-Ec.oriV-RK2 Agro transformation B-ARGtu.left border Barker, R. F. etal (1983) Plant Mol Biol. 2:335-350 Plant selectable marker expression Promoter with intron and McDowell et al. (1996) caSSette 5'UTR of Arabidopsis act 7 Plant Physiol. 111: 699-711. 5' UTR of Arabidopsis act 7 gene Intron in SUTR of AtAct 7 Transit peptide region of Klee, H. J. etal (1987) Arabidopsis EPSPS MGG 210: 437-442 Synthetic CP4 coding region with dicot preferred codon usage A 3' UTR of the nopaline U.S. Pat. No. 5,858,742 synthase gene of Agrobacterium tumefaciens Tiplasmid Plant gene of interest expression Promoter for 35S RNA from U.S. Pat. No. 5,322,938 caSSette CaMV containing a duplication of the -90 to -350 region US 2008/0148432 A1 Jun. 19, 2008

TABLE 6-continued

Function Element Reference Gene of interest insertion site Cotton E63' end GenBank accession U3OSO8

0069 Primers for PCR amplification of protein coding bardment, a suspension of gold particles is prepared onto nucleotides of recombinant DNA were designed at or near the which the desired recombinant DNA expression cassettes are start and stop codons of the coding sequence, in order to precipitated. DNA is introduced into maize cells as described eliminate most of the 5' and 3' untranslated regions. Each in U.S. Pat. Nos. 5,550,318 and 6,399,861 using the electric recombinant DNA coding for a protein identified in Table 1 discharge particle acceleration gene delivery device. Follow was amplified by PCR prior to insertion into the insertion site ing microprojectile bombardment, tissue is cultured in the of one of the base vectors as referenced in Table 1. dark at 27 degrees C. Additional transformation methods and materials for making transgenic plants of this invention, for EXAMPLE 2 example, various media and recipient target cells, transfor mation of immature embryos and Subsequence regeneration Corn Transformation of fertile transgenic plants are disclosed in U.S. Pat. Nos. 0070 This example illustrates plant cell transformation 6,194.636 and 6.232,526 and U.S. patent application Ser. No. methods useful in producing transgenic corn plant cells, 09/757,089, which are incorporated herein by reference. plants, seeds and pollen of this invention and the production 0074 To regenerate transgenic corn plants a callus of and identification of transgenic corn plants and seed with an transgenic plant cells resulting from transformation is placed enhanced trait, i.e. enhanced water use efficiency, enhanced on media to initiate shoot development in plantlets which are cold tolerance, increased yield, enhanced nitrogen use effi transferred to potting soil for initial growth in a growth cham ciency, enhanced seed protein and enhanced seed oil. Plasmid ber at 26 degrees C. followed by a mist bench before trans vectors were prepared by cloning DNA identified in Table 1 in planting to 5 inch pots where plants are grown to maturity. the identified base vectors for use in corn transformation of The regenerated plants are self fertilized and seedisharvested cornplant cells to produce transgenic corn plants and progeny for use in one or more methods to select seed, seedlings or plants, seed and pollen. progeny second generation transgenic plants (R2 plants) or 0071. For Agrobacterium-mediated transformation of hybrids, e.g. by selecting transgenic plants exhibiting an corn embryo cells corn plants of a readily transformable line enhanced trait as compared to a control plant. (designated LH59) is grown in the greenhouse and ears har 0075 Transgenic corn plant cells are transformed with vested when the embryos are 1.5 to 2.0 mm in length. Ears are recombinant DNA from each of the genes identified in Table Surface sterilized by spraying or soaking the ears in 80% 1. Progeny transgenic plants and seed of the transformed ethanol, followed by air drying. Immature embryos are iso plant cells are screened for enhanced water use efficiency, lated from individual kernels on surface sterilized ears. Prior enhanced cold tolerance, increased yield, enhanced nitrogen to inoculation of maize cells, Agrobacterium cells are grown use efficiency, enhanced seed protein and enhanced seed oil overnight at room temperature. Immature maize embryo cells as reported in Example 5. are inoculated with Agrobacterium shortly after excision, and incubated at room temperature with Agrobacterium for 5-20 EXAMPLE 3 minutes. Immature embryo plant cells are then co-cultured with Agrobacterium for 1 to 3 days at 23°C. in the dark. Soybean Transformation Co-cultured embryos are transferred to selection media and 0076. This example illustrates plant transformation useful cultured for approximately two weeks to allow embryogenic in producing the transgenic soybean plants of this invention callus to develop. Embryogenic callus is transferred to culture and the production and identification of transgenic Seed for medium containing 100 mg/L paromomycin and Subcultured transgenic soybean having enhanced water use efficiency, at about two week intervals. Transformed plant cells are enhanced cold tolerance, increased yield, enhanced nitrogen recovered 6 to 8 weeks after initiation of selection. use efficiency, enhanced seed protein and enhanced seed oil. 0072 For Agrobacterium-mediated transformation of 0077. For Agrobacterium mediated transformation, soy maize callus immature embryos are cultured for approxi bean seeds are germinated overnight and the meristem mately 8-21 days after excision to allow callus to develop. explants excised. The meristems and the explants are placed Callus is then incubated for about 30 minutes at room tem in a wounding vessel. Soybean explants and induced Agro perature with the Agrobacterium suspension, followed by bacterium cells from a strain containing plasmid DNA with removal of the liquid by aspiration. The callus and Agrobac the gene of interest cassette and a plant selectable marker terium are co-cultured without selection for 3-6 days fol cassette are mixed no later than 14 hours from the time of lowed by selection on paromomycin for approximately 6 initiation of seed germination and wounded using Sonication. weeks, with biweekly transfers to fresh media, and paromo Following wounding, explants are placed in co-culture for 2-5 mycin resistant callus identified as containing the recombi days at which point they are transferred to selection media for nant DNA in an expression cassette. 6-8 weeks to allow selection and growth of transgenic shoots. 0073 For transformation by microprojectile bombard Trait positive shoots are harvested approximately 6-8 weeks ment immature maize embryos are isolated and cultured 3-4 and placed into selective rooting media for 2-3 weeks. Shoots days prior to bombardment. Prior to microprojectile bom producing roots are transferred to the greenhouse and potted US 2008/0148432 A1 Jun. 19, 2008 26 in soil. Shoots that remain healthy on selection, but do not 166 and homologs of SEQ ID NO: 1482 through 52767 is produce roots are transferred to non-selective rooting media identified in Table 2. The source organism for each homolog for an additional two weeks. Roots from any shoots that is found in the Sequence Listing. produce roots off selection are tested for expression of the plant selectable marker before they are transferred to the EXAMPLE 5 greenhouse and potted in soil. Additionally, a DNA construct can be transferred into the genome of a Soybean cell by Selection of Transgenic Plants with Enhanced Agro particle bombardment and the cell regenerated into a fertile nomic Trait(s) soybean plant as described in U.S. Pat. No. 5,015,580, herein I0083. This example illustrates identification of plant cells incorporated by reference. of the invention by Screening derived plants and seeds for 0078 Transgenic soybean plant cells are transformed with enhanced trait. Transgenic corn seed and plants with recom recombinant DNA from each of the genes identified in Table binant DNA identified in Table 1 were prepared by plant cells 1. Progeny transgenic plants and seed of the transformed transformed with DNA that was stably integrated into the plant cells are screened for enhanced water use efficiency, genome of the corn cell. The transgenic seed, plantlets and enhanced cold tolerance, increased yield, enhanced nitrogen progeny plants were selected using the methods that measure use efficiency, enhanced seed protein and enhanced seed oil Transgenic corn plant cells were transformed with recombi as reported in Example 5. nant DNA from each of the genes identified in Table 1. Prog eny transgenic plants and seed of the transformed plant cells EXAMPLE 4 were screened for enhanced water use efficiency, enhanced cold tolerance, increased yield, enhanced nitrogen use effi Homolog Identification ciency, enhanced seed protein and enhanced seed oil as com 0079. This example illustrates the identification of pared to control plants. homologs of proteins encoded by the DNA identified in Table 1 which is used to provide transgenic seed and plants having A. Selection for Enhanced Nitrogen Use Efficiency enhanced agronomic traits. From the sequence of the I0084. The physiological efficacy of transgenic corn plants homologs, homologous DNA sequence can be identified for (tested as hybrids) can be tested for nitrogen use efficiency preparing additional transgenic seeds and plants of this inven (NUE) traits in a high-throughput nitrogen (N) selection tion with enhanced agronomic traits. method. The collected data are compared to the measure 0080. An All Protein Database' was constructed of ments from wildtype controls using a statistical model to known protein sequences using a proprietary sequence data determine if the changes are due to the transgene. Raw data base and the National Center for Biotechnology Information were analyzed by SAS software. Results shown herein are the (NCBI) non-redundantamino acid database (nraa). For each comparison of transgenic plants relative to the wildtype con organism from which a polynucleotide sequence provided trols. herein was obtained, an "Organism Protein Database' was constructed of known protein sequences of the organism; it is (1) Media Preparation for Planting a NUE Protocol a Subset of the All Protein Database based oil the NCBI taxonomy ID for the organism. I0085 Planting materials used: Metro Mix 200 (vendor: 0081. The All Protein Database was queried using amino Hummert) Cat. # 10-0325, Scotts Micro Max Nutrients (ven acid sequences provided herein as SEQID NO: 142 through dor: Hummert) Cat. # 07-6330, OS 4/3"x37/8" pots (vendor: SEQID NO:1482 using NCBI “blastp' program with E-value Hummert) Cat. # 16-1415, OS trays (vendor: Hummert) Cat. cutoff of 1e-8. Up to 1000 top hits were kept, and separated it 16-1515, Hoagland's macronutrients solution, Plastic 5" by organism names. For each organism other than that of the stakes (vendor: Hummert) yellow Cat. ii 49-1569, white Cat. query sequence, a list was kept for hits from the query organ # 49-1505, Labels with numbers indicating material con ism itself with a more significant E-value than the best hit of tained in pots. Fill 500 pots to rim with Metro Mix 200 to a the organism. The list contains likely duplicated genes of the weight of ~140 g/pot. Pots are filled uniformly by using a polynucleotides provided herein, and is referred to as the balancer. Add 0.4 g of Micro Max nutrients to each pot. Stir Core List. Another list was kept for all the hits from each ingredients with spatula to a depth of 3 inches while prevent organism, sorted by E-value, and referred to as the Hit List. ing material loss. 0082. The Organism Protein Database was queried using polypeptide sequences provided herein as SEQ ID NO: 142 (2) Planting a NUE Selection in the Greenhouse through SEQID NO:1482 using NCBI “blastp'program with I0086 (a) Seed Germination. Each pot is lightly altered E-value cutoff of 1e-4. Up to 1000 top hits were kept. A twice using reverse osmosis purified water. The first watering BLAST searchable database was constructed based on these is scheduled to occur just before planting; and the second hits, and is referred to as “SubDB'. SubDB was queried with watering, after the seed has been planted in the pot. Ten Seeds each sequence in the Hit List using NCBI “blastp' program of each entry (1 seed per pot) are planted to select eight with E-value cutoff of 1e-8. The hit with the best E-value was healthy uniform seedlings. Additional wild type controls are compared with the Core List from the corresponding organ planted for use as border rows. Alternatively, 15 seeds of each ism. The hit is deemed a likely ortholog if it belongs to the entry (1 seed per pot) are planted to select 12 healthy uniform Core List, otherwise it is deemed not a likely ortholog and seedlings (this larger number of plantings is used for the there is no further search of sequences in the Hit List for the second, or confirmation, planting). Place pots on each of the same organism. Homologs from a large number of distinct 12 shelves in the Conviron growth chamber for seven days. organisms were identified and are reported by amino acid This is done to allow more uniform germination and early sequences of SEQID NO: 1483 through SEQID NO:52767. seedling growth. The following growth chamber settings are These relationship of proteins of SEQ ID NO: 742 through 25°C./day and 22°C./night, 14 hours light and ten hours dark, US 2008/0148432 A1 Jun. 19, 2008 27 humidity ~80%, and light intensity ~350 mol/m/s (at pot base) with approximately equal spacing between them. The level). Watering is done via capillary matting similar to green average of the three measurements was taken from the SPAD house benches with duration often minutes three times a day. machine. 0087 (b) Seedling transfer After seven days, the best 0091 Leaf fresh mass is recorded for an excised V6 leaf, eight or 12 seedlings for the first or confirmation pass runs, the leaf is placed into a paper bag. The paper bags containing respectively, are chosen and transferred to greenhouse the leaves are then placed into a forced air oven at 80°C. for benches. The pots are spaced eight inches apart (center to 3 days. After 3 days, the paper bags are removed from the center) and are positioned on the benches using the spacing oven and the leaf dry mass measurements are taken. patterns printed on the capillary matting. The Vattex matting 0092. From the collected data, two derived measurements creates a 384-position grid, randomizing all range, row com are made: (1) Leaf chlorophyll area (LCA), which is a product binations. Additional pots of controls are placed along the of V6 relative chlorophyll content and its leaf area (relative outside of the experimental block to reduce border effects. units). Leaf chlorophyll area-leaf chlorophyll X leaf area. 0088 Plants are allowed to grow for 28 days under the low This parameter gives an indication of the spread of chloro N run or for 23 days under the high N run. The macronutrients phyll over the entire leaf area; (2) specific leafarea (LSA) is are dispensed in the form of a macronutrient solution (see calculated as the ratio of V6 leaf area to its dry mass (cm/g composition below) containing precise amounts of N added dry mass), a parameter also recognized as a measure of NUE. (2 mMNHNO, for limiting N selection and 20 mMNHNO, The data are shown in Table 8. for high N selection runs). Each pot is manually dispensed 100 ml of nutrient solution three times a week on alternate Nitrogen Use Field Efficacy Assay days starting at eight and ten days after planting for high N and low N runs, respectively. On the day of nutrient applica 0093 Level I. Transgenic plants provided by the present tion, two 20 min waterings at 05:00 and 13:00 are skipped. invention are planted in field without any nitrogen source The Vattex matting should be changed every third run to avoid being applied. Transgenic plants and control plants are Naccumulation and buildup of root matter. Table 7 shows the grouped by genotype and construct with controls arranged amount of nutrients in the nutrient solution for either the low randomly within genotype blocks. Each type of transgenic plants are tested by 3 replications and across 5 locations. or high nitrogen selection. Nitrogen levels in the fields are analyzed in early April pre planting by collecting 30 sample soil cores from 0-24" and 24 TABLE 7 to 48" soil layer. Soil samples are analyzed for nitrate-nitro 2 mM NHNO 20 mMNHNO gen, phosphorus (P), Potassium (K), organic matter and pH to (Low Nitrogen (high provide baseline values. P. K and micronutrients are applied Growth Nitrogen Growth Condition, Condition, based upon Soil test recommendations. Low N) High N) 0094 Level II. Transgenic plants provided by the present Nutrient Stock mLL mL/L invention are planted in field with three levels of nitrogen (N) 1 MNHNO 2 2O fertilizer being applied, i.e. low level (ON), medium level (80 1 M KHPO. O.S O.S lbfac) and high level (180 lbfac). Liquid 28% or 32% UAN 1 M MgSO4·7H2O 2 2 (Urea, Ammonium Nitrogen) are used as the N source and 1 M CaCl, 2.5 2.5 apply by broadcast boom and incorporate with a field culti 1 MK2SO 1 1 vator with rearrolling basket in the same direction as intended Note: crop rows. Although there is no Napplied to the ON treatment Adjust pH to 5.6 with HCl or KOH the soil should still be disturbed in the same fashion as the treated area. Transgenic plants and control plants are grouped 0089 (c) Harvest Measurements and Data Collection— by genotype and construct with controls arranged randomly After 28 days of plant growth for low N runs and 23 days of within genotype blocks. Each type of transgenic plants is plant growth for high N runs, the following measurements are tested by 3 replications and across 4 locations. Nitrogen levels taken (phenocodes in parentheses): total shoot fresh mass (g) in the fields are analyzed in early April pre-planting by col (SFM) measured by Sartorius electronic balance, V6 leaf lecting 30 sample soil cores from 0-24" and 24 to 48" soil chlorophyll measured by Minolta SPAD meter (relative units) layer. Soil samples are analyzed for nitrate-nitrogen, phos (LC). V6 leafarea (cm) (LA) measured by a Li-Corleafarea phorus (P), Potassium (K), organic matter and pH to provide meter, V6 leaf fresh mass (g) (LFM) measured by Sartorius baseline values. P. K and micronutrients are applied based electronic balance, and V6 leaf dry mass (g) (LDM) measured upon soil test recommendations. by Sartorius electronic balance. Raw data were analyzed by SAS software. Results shown are the comparison of trans B. Selection for Increased Yield genic plants relative to the wildtype controls. 0090. To take a leaf reading, samples were excised from 0.095 Many transgenic plants of this invention exhibit the V6 leaf. Since chlorophyll meter readings of corn leaves improved yield as compared to a control plant. Improved are affected by the part of the leaf and the position of the leaf yield can result from enhanced seed sink potential, i.e. the on the plant that is sampled, SPAD meter readings were done number and size of endosperm cells or kernels and/or on leaf six of the plants. Three measurements per leaf were enhanced sink strength, i.e. the rate of Starch biosynthesis. taken, of which the first reading was taken from a point Sink potential can be established very early during kernel one-half the distance between the leaf tip and the collar and development, as endosperm cell number and size are deter halfway from the leaf margin to the midrib while two were mined within the first few days after pollination. taken toward the leaf tip. The measurements were restricted in 0096. Much of the increase in cornyield of the past several the area from /2 to 3/4 of the total length of the leaf (from the decades has resulted from an increase in planting density. US 2008/0148432 A1 Jun. 19, 2008 28

During that period, cornyield has been increasing at a rate of ment include Source capacity (biomass), source output (Su 2.1 bushels/acre?year, but the planting density has increased crose and photosynthesis), sink components (kernel size, ear at a rate of 250 plants/acre?year. A characteristic of modern size, starch in the seed), development (light response, height, hybrid corn is the ability of these varieties to be planted at density tolerance), maturity, early flowering trait and physi high density. Many studies have shown that a higher than ological responses to high density planting, for example at current planting density should result in more biomass pro 45,000 plants per acre, for example as illustrated in Table 10 duction, but current germplasm does not perform. well at and 11.

TABLE 8

Timing Evaluation Description connents V2-3 Early stand Can be taken any time after germination and prior to removal of any plants. Pollen shed GDU to 50% shed GDU to 50% plants shedding 50% tassel. Silking GDU to 50% silk GDU to 50% plants showing silks. Maturity Plant height Height from soil surface to 10 plants per plot-Yield flag leaf attachment (inches). team assistance Maturity Ear height Height from soil surface to 10 plants per plot-Yield primary ear attachment node. team assistance Maturity Leaves above ear visual scores: erect, size, rolling Maturity Tassel size Visual scores + - vs. WT Pre-Harwest Final Stand Final stand count prior to harvest, exclude tillers Pre-Harwest Stalk lodging No. of stalks broken below the primary ear attachment. Exclude leaning tillers Pre-Harwest Root lodging No. of stalks leaning >45° angle from perpendicular. Pre-Harwest Stay green After physiological maturity and when differences among genotypes are evident: Scale 1 (90-100% tissue green)-9 (0-19% tissue green). Harvest Grain Yield Grain yield plot (Shell weight) these higher densities. One approach to increasing yield is to increase harvest index (HI), the proportion of biomass that is TABLE 9 allocated to the kernel compared to total biomass, in high density plantings. Timing Evaluation Description V8-V12 Chlorophyll 0097 Effective yield selection of enhanced yielding trans V12-VT Ear leafarea genic corn events uses hybrid progeny of the transgenic event W15-1SDAP Chl fluorescence over multiple locations with plants grown under optimal pro W15-1SDAP CER duction management practices, and maximum pest control. A 1S-2S DAP Carbohydrates Sucrose, starch Pre-Harwest 1st internode diameter useful target for improved yield is a 5% to 10% increase in Pre-Harwest Base 3 internode diameter yield as compared to yield produced by plants grown from Pre-Harwest Ear internode diameter seed for a control plant. Selection methods may be applied in Maturity Ear traits diameter, length, kernel multiple and diverse geographic locations, for example up to number, kernel weight 16 or more locations, over one or more plating seasons, for example at least two planting seasons to statistically distin guish yield improvement from natural environmental effects. I0098 Electron transport rates (ETR) and CO, exchange It is to plant multiple transgenic plants, positive and negative rates (CER): ETR and CER are measured with Li6400LCF control plants, and pollinator plants in Standard plots, for (Licor, Lincoln, Nebr.) around V9-R1 stages. Leaf chloro example 2 row plots, 20 feet long by 5 feet wide with 30 phyll fluorescence is a quick way to monitor the source activ inches distance between rows and a 3 foot alley between ity and is reported to be highly correlated with CO assimi ranges. Transgenic events can be grouped by recombinant lation under varies conditions (Photosyn Research, 37: DNA constructs with groups randomly placed in the field. A 89-102). The youngest fully expanded leafor 2 leaves above pollinator plot of a high quality corn line is planted for every the ear leaf is measured with actinic light 1500 (with 10% two plots to allow open pollination when using male sterile blue light) micromol mis", 28°C., CO levels 450 ppm. transgenic events. A useful planting density is about 30,000 Ten plants are measured in each event. There were 2 readings plants/acre. High planting density is greater than 30,000 for each plant. plants/acre, preferably about 40,000 plants/acre, more pref (0099. A hand-held chlorophyll meter SPAD-502 (Mi erably about 42,000 plants/acre, most preferably about nolta—Japan) is used to measure the total chlorophyll level 45,000 plants/acre. Surrogate indicators for yield improve on live transgenic plants and the wild type counterparts a. US 2008/0148432 A1 Jun. 19, 2008 29

Three trifoliates from each plant are analyzed, and each tri water cycles on plants over a total period of 15 days after an foliate were analyzed three times. Then 9 data points are initial stress free growth period of 11 days. Each cycle con averaged to obtain the chlorophyll level. The number of ana sists of 5 days, with no water being applied for the first four lyzed plants of each genotype ranges from 5 to 8. days and a water quenching on the 5th day of the cycle. The 0100 When selecting for yield improvement a useful sta primary phenotypes analyzed by the selection method are the tistical measurement approach comprises three components, changes in plant growth rate as determined by height and i.e. modeling spatial autocorrelation of the test field sepa biomass during a vegetative drought treatment. The hydration rately for each location, adjusting traits of recombinant DNA status of the shoot tissues following the drought is also mea events for spatial dependence for each location, and conduct sured. The plant height are measured at three tine points. The ing an across location analysis. The first step in modeling first is taken just prior to the onset drought when the plant is spatial autocorrelation is estimating the covariance param 11 days old, which is the shoot initial height (SIH). The plant eters of the semivariogram. A spherical covariance model is height is also measured halfway throughout the drought/re assumed to model the spatial autocorrelation. Because of the size and nature of the trial, it is likely that the spatial autocor water regimen, on day 18 after planting, to give rise to the relation may change. Therefore, anisotropy is also assumed shoot mid-drought height (SMH). Upon the completion of the along with spherical covariance structure. The following set final drought cycle on day 26 after planting, the shoot portion of equations describes the statistical form of the anisotropic of the plant is harvested and measured for a final height, spherical covariance model. which is the shoot wilt height (SWH) and also measured for shoot wilted biomass (SWM). The shoot is placed in water at 40 degree Celsius in the dark. Three days later, the shoot is 2 3 1 3 weighted to give rise to the shoot turgid weight (STM). After Ch; 0) = vich =0) + or 1-5 high ?ch - 1), drying in an oven for four days, the shoots are weighted for shoot dry biomass (SDM). The shoot average height (SAH) is the mean plant height across the 3 height measurements. The 0101 where I() is the indicator function, h=yx+y, and procedure described above may be adjusted for +/--one day for each step given the situation. 0104. To correct for slight differences between plants, a size corrected growth value is derived from SIH and SWH. where S=(x, y) are the spatial coordinates of one location This is the Relative Growth Rate (RGR). Relative Growth and S(x, y) are the spatial coordinates of the second Rate (RGR) is calculated for each shoot using the formula location. There are 5 covariance parameters, 0=(V, of, p. (), RGR 96=(SWH-SIH)/((SWH+SIH)/2)* 100. Relative co), where v is the nugget effect, of is the partial sill, p is a water content (RWC) is a measurement of how much (%) of rotation in degrees clockwise from north, (), is a scaling the plant was water at harvest. Water Content (RwC) is cal parameter for the minor axis and (), is a scaling parameter for culated for each shoot using the formula RWC 96 (SWM the major axis of an anisotropical ellipse of equal covariance. SDM)/(STM-SDM)*100). Fully watered corn plants of this The five covariance parameters that defines the spatial trend age run around 98% RWC. will then be estimated by using data from heavily replicated pollinator plots via restricted maximum likelihood approach. D. Selection for Growth Under Cold Stress In a multi-location field trial, spatial trend are modeled sepa 0105 (1) Cold germination assay Three sets of seeds are rately for each location. used for the assay. The first set consists of positive transgenic 0102. After obtaining the variance parameters of the events (F1 hybrid) where the genes of the present invention model, a variance-covariance structure is generated for the are expressed in the seed. The second seed set is nontrans data set to be analyzed. This variance-covariance structure genic, wild-type negative control made from the same geno contains spatial information required to adjust yield data for type as the transgenic events. The third set consisted of two spatial dependence. In this case, a nested model that best cold tolerant and one cold sensitive commercial checklines of represents the treatment and experimental design of the study corn. All seeds are treated with a fungicide “Captan’ (MAE is used along with the variance-covariance structure to adjust STROR 80DF Fungicide, Arvesta Corporation, San Fran the yield data. During this process the nursery or the seed cisco, Calif., USA). 0.43 mL Captain is applied per 45 g of batch effects can also be modeled and estimated to adjust the corn seeds by mixing it well and drying the fungicide prior to yields for any yield parity caused by seed batch differences. the experiment. After spatially adjusted data from different locations are gen erated, all adjusted data is combined and analyzed assuming 0106 Corn kernels are placed embryo side down on blot locations as replications. In this analysis, intra and inter ter paper within an individual cell (8.9x8.9 cm) of a germi location variances are combined to estimate the standard error nation tray (54x36 cm). Ten seeds from an event are placed of yield from transgenic plants and control plants. Relative into one cell of the germination tray. Each tray can hold 21 mean comparisons are used to indicate statistically significant transgenic events and 3 replicates of wildtype (LH244SDms+ yield improvements. LH59), which is randomized in a complete block design. For every event there are five replications (five trays). The trays C. Selection for Enhanced Water Use Efficiency (WUE) are placed at 9.7C for 24 days (no light) in a Convrion growth chamber (Conviron Model PGV36, Controlled Environ 0103) Described in this example is a high-throughput ments, Winnipeg, Canada). Two hundred and fifty milliliters method for greenhouse selection of transgenic corn plants to of deionized water are added to each germination tray. Ger wild type corn plants (tested as inbreds or hybrids) for water mination counts are taken 10th, 11th, 12th, 13th, 14th, 17th, use efficiency. This selection process imposes 3 drought/re 19th, 21st, and 24th day after start date of the experiment. US 2008/0148432 A1 Jun. 19, 2008 30

Seeds are considered germinated if the emerged radical size is the same genotype as the transgenic events. The third seed set 1 cm. From the germination counts germination index is consists of two cold tolerant and two cold sensitive commer calculated. cial check lines of corn. All seeds are treated with a fungicide 0107 The germination index iv calculated as per. “Captan', (3a,4,7a-tetrahydro-2-(trichloromethly)thio)-1H isoindole-1,3-(2H)-dione, Drex Chemical Co. Memphis, Germination index=(X(T+1-n, IP-P, ))/T Tenn.). Captain (0.43 mL) was applied per 45g of corn seeds 0108. Where T is the total number of days for which the by mixing it well and drying the fungicide prior to the experi germination assay is performed. The number of days after ment. planting is defined by n. “i' indicated the number of times the 0114 Seeds are grown in germination paper for the early germination had been counted, including the current day. Pis seedling growth assay. Three 12" x 18" pieces of germination the percentage of seeds germinated during any given rating. paper (Anchor Paper iSD7606) are used for each entry in the Statistical differences are calculated between transgenic test (three repetitions per transgenic event). The papers are events and wild type control. After statistical analysis, the wetted in a solution of 0.5% KNO, and 0.1% Thyram. events that show a statistical significance at the p level of less 0115 For each paper fifteen seeds are placed on the line than 0.1 relative to wild-type controls will advance to a sec evenly spaced down the length of the paper. The fifteen seeds ondary cold selection. The secondary cold Screen is con are positioned on the paper Such that the radical would grow ducted in the same manner of the primary selection only downward, for example longer distance to the paper's edge. increasing the number of repetitions to ten. Statistical analy The wet paper is rolled up starting from one of the short ends. sis of the data from the secondary selection is conducted to The paper is rolled evenly and tight enough to hold the seeds identify the events that show a statistical significance at the p in place. The roll is secured into place with two large paper level of less than 0.05 relative to wild-type controls. clips, one at the top and one at the bottom. The rolls are 0109 (2) Cold Shock assay The experimental set-up for incubated in a growth chamber at 23°C. for three days in a the cold shockassay is the same as described in the above cold randomized complete block design within an appropriate germination assay except seeds were grown in potted media container. The chamber is set for 65% humidity with no light for the cold shock assay. cycle. For the cold stress treatment the rolls are then incubated 0110. The desired numbers of 2.5" square plastic pots are in a growth chamber at 12°C. for twelve days. The chamber placed on flats (n=32, 4x8). Pots were filled with Metro Mix is set for 65% humidity with no light cycle. 200 soil-less media containing 19:6:12 fertilizer (6 lbs/cubic 0116. After the cold treatment the germination papers are yard) (Metro Mix, Pots and Flat are obtained from Hummert unrolled and the seeds that did not germinate are discarded. International, Earth City, Mo.). After planting seeds, pots are The lengths of the radical and coleoptile for each seed are placed in a growth chamber set at 23°C., relative humidity of measured through an automated imaging program that auto 65% with 12 hour day and night photoperiod (300 uE/m2 matically collects and processes the images. The imaging min). Planted seeds are watered for 20 minute every other day program automatically measures the shoot length, root by sub-irrigation and flats were rotated every third day in a length, and whole seedling length of every individual seed growth chamber for growing corn seedlings. ling and then calculates the average of each roll. I0111. On the 10" day afterplanting the transgenic positive 0117. After statistical analysis, the events that show a sta and wild-type negative (WT) plants are positioned in flats in tistical significance at the p level of less than 0.1 relative to an alternating pattern. Chlorophyll fluorescence of plants is wild-type controls will advance to a secondary cold selection. measured on the 10" day during the dark period of growth by The secondary cold selection is conducted in the same man using a PAM-2000 portable fluorometer as per the manufac ner of the primary selection only increasing the number of turer's instructions (Walz, Germany). After chlorophyll mea repetitions to five. Statistical analysis of the data from the Surements, leaf samples from each event are collected for secondary selection is conducted to identify the events that confirming the expression of genes of the present invention. show a statistical significance at the p level of less than 0.05 For expression analysis six V1 leaf tips from each selection relative to wild-type controls. are randomly harvested. The flats are moved to a growth chamber set at 5° C. All other conditions Such as humidity, 4. Cold Field Efficacy Trial day/night cycle and light intensity are held constant in the 0118. This example sets forth a cold field efficacy trial to growth chamber. The flats are sub-irrigated every day after identify gene constructs that confer enhanced cold vigor at transfer to the cold temperature. On the 4" day chlorophyll germination and early seedling growth under early spring fluorescence is measured. Plants are transferred to normal planting field conditions in conventional-till and simulated growth conditions after six days of cold shock treatment and no-till environments. Seeds are planted into the ground allowed to recover for the next three days. During this recov around two weeks before local farmers are beginning to plant ery period the length of the V3 leaf is measured on the 1 and corn so that a significant cold stress is exerted onto the crop, 3" days. After two days of recovery V2 leaf damage is deter named as cold treatment. Seeds also are planted under local mined visually by estimating percent of green V2 leaf. optimal planting conditions such that the crop has little or no 0112 Statistical differences in V3 leaf growth, V2 leaf exposure to cold condition, named as normal treatment. The necrosis and fluorescence during pre-shock and cold shock cold field efficacy trials are carried out in five locations, can be used for estimation of cold shock damage on corn including Glyndon Minn., Mason Mich., Monmouth Ill., plants. Dayton Iowa, Mystic Conn. At each location, seeds are 0113 (3) Early seedling growth assay Three sets of planted under both cold and normal conditions with 3 repeti seeds are used for the experiment. The first set consists of tions per treatment, 20 kernels per row and single row per positive transgenic events (F1 hybrid) where the genes of the plot. Seeds are planted 1.5 to 2 inch deep into soil to avoid present invention are expressed in the seed. The second seed muddy conditions. Two temperature monitors are set up at set is nontransgenic, wild-type negative control made from each location to monitor both air and soil temperature daily. US 2008/0148432 A1 Jun. 19, 2008

0119 Seed emergence is defined as the point when the EXAMPLE 6 growing shoot breaks the soil surface. The number of Consensus Sequence emerged seedling in each plot is counted everyday from the day the earliest plot begins to emerge until no significant 0.122 This example illustrates the identification of con changes in emergence occur. In addition, for each planting sensus amino acid sequence for the proteins and homologs encoded by DNA that is used to prepare the transgenic seed date, the latest date when emergence is 0 in all plots is also and plants of this invention having enhanced agronomic recorded. Seedling vigor is also rated at V3-V4 stage before traits. the average of corn plant height reaches 10 inches, with I0123 ClustalW program was selected for multiple 1=excellent early growth, 5–Average growth and 9-poor sequence alignments of the amino acid sequence of SEQID growth. Days to 50% emergence, maximum percent emer NO: 1205 and its 12 homologs. Three major factors affecting gence and seedling vigor are calculated using SAS Software the sequence alignments dramatically are (1) protein weight for the data within each location or across all locations. matrices; (2) gap open penalty; (3) gap extension penalty. E. Screens for Transgenic Plant Seeds with Increased Protein Protein weight matrices available for ClustalW program and/or Oil Levels include Blosum, Pam and Gonnet series. Those parameters 0120. This example sets forth a high-throughput selection with gap open penalty and gap extension penalty were exten for identifying plant seeds with improvement in seed compo sively tested. On the basis of the test results, Blosum weight sition using the Infratec 1200 series Grain Analyzer, which is matrix, gap open penalty of 10 and gap extension penalty of 1 a near-infrared transmittance spectrometer used to determine were chosen for multiple sequence alignment. FIG. 2 shows the composition of a bulk seed sample. Near infrared analysis the sequences of SEQ ID NO: 1205, its homologs and the is a non-destructive, high-throughput method that can ana consensus sequence (SEQ ID NO: 52803) at the end. The lyZe multiple traits in a single sample scan. An NIR calibra symbols for consensus sequence are (1) uppercase letters for tion for the analytes of interest is used to predict the values of 100% identity in all positions of multiple sequence alignment an unknown sample. The NIR spectrum is obtained for the output; (2) lowercase letters for >=70% identity: symbol; (3) sample and compared to the calibration using a complex “X” indicated <70% identity; (4) dashes "- meaning that chemometric Software package that provides a predicted val gaps were in >=70% sequences. ues as well as information on how well the sample fits in the 0.124. The consensus amino acid sequence can be used to calibration. identify DNA corresponding to the full scope of this invention that is useful in providing transgenic plants, for example corn 0121 Infratec Model 1221, 1225, or 1227 with transport and soybean plants with enhanced agronomic traits, for module by Foss North America is used with cuvette, item if example improved nitrogen use efficiency, improved yield, 1000-4033, Foss North America or for small samples with small cell cuvette, Foss standard cuvette modified by Leon improved water use efficiency and/or improved growth under Girard Co. Corn and soy check samples of varying composi cold stress, due to the expression in the plants of DNA encod tion maintained in check cell cuvettes are Supplied by Leon ing a protein with amino acid sequence identical to the con Girard Co. NIT collection software is provided by Maximum sensus amino acid sequence. Consulting Inc. Software. Calculations are performed auto EXAMPLE 7 matically by the software. Seed samples are received in pack Identification of Amino Acid Domain by Pfam ets or containers with barcode labels from the customer. The Analysis seed is poured into the cuvettes and analyzed as received. 0.125. The amino acid sequence of the expressed proteins TABLE 10 that were shown to be associated with an enhanced trait were analyzed for Pfam protein family against the current Pfam Typical sample(s): Whole grain corn and soybean seeds collection of multiple sequence alignments and hidden Analytical time to run method: Less than 0.75 min per sample Total elapsed time per run: 1.5 minute per sample Markov models using the HMMER software in the appended Typical and minimum sample size: Corn typical: 50 cc, minimum 30 cc computer listing. The Pfam protein families for the proteins of Soybean typical: 50 cc. SEQ ID NO: 742 through 1482 are shown in Table 11. The minimum 5 cc Typical analytical range: Determined in part by the specific Hidden Markov model databases for the identified patent calibration. families are also in the appended computer listing allowing Corn - moisture 5-15%, oil 5-20%, identification of other homologous proteins and their cognate protein 5-30%, starch 50-75%, encoding DNA to enable the full breadth of the invention for and density 1.0-1.3%. a person of ordinary skill in the art. Certain proteins are Soybean - moisture 5-15%, oil 15-25%, identified by a single Pfam domain and others by multiple and protein 35-50%. Pfam domains. For instance, the protein with amino acids of SEQ ID NO: 817 is characterized by two Pfam domains, i.e. GTP EFTU, GTP EFTU D2 and GTP EFTU D3. TABLE 11

PEP SEQ NUC ID SEQ NO ID NO GENEID Pfam domain name begin stop score E-value

816 75 PHEOOOOO35 61 bZIP 1 15 79 41.9 2.OOE-09 816 75 PHEOOOOO35 61 bZIP 2 15 69 42.8 1.1OE-09 US 2008/0148432 A1 Jun. 19, 2008 32

TABLE 11-continued

PEP SEQ NUC ID SEQ NO ID NO GENEID Pfam domain name begin stop SCO 817 76 EOOOOO36 62 GTP EFTU 5 227 359.5 -105 817 76 EOOOOO36 62 GTP EFTU D2 248 315 91.9 24 817 76 EOOOOO36 62 GTP EFTU D3 322 430 2O7.6 818 77 EOOOO 30 220 acalin 185 318 84.5 1177 436 EOOOO 32 222 PCI 63 159 44.2 819 78 EOOOO 34 224 LEA 5 1 113 151.2 820 79 EOOOO 35 225 PP2C 39 281 226.2 821 8O EOOOO 36 226 Hist deacetyl 31 344 579.9 -171 822 81 EOOOO 39 229 peroxidase 19 227 239.9 823 82 EOOOO 41. 231 184 295 -0.9 824 83 EOOOO 42 232 11 179 28O.2 825 84 EOOOO 46 236 13 174 335 826 85 EOOOO 48 238 12 267 323.2 826 85 EOOOO 48 238 Pkinase Tyr 12 26S 70.9 826 85 EOOOO 48 238 NAF 310 369 116.1 827 86 EOOOO 49 239 AP2 29 92 128.5 828 87 EOOOO 50 240 Lectin legB 23 212 828 87 EOOOO 50 240 Lectin legA 221 26S 37.6 828 87 EOOOO 50 240 Pkinase 344 496 -11.1 829 88 EOOOO 89 282 HSP20 49 152 169.6 830 89 E0000200 293 22 317 296.5 831 90 E0000211. 304 Pkinase 30 288 384.1 -112 832 91 E0000213 306 Pkinase 64 322 348.7 -102 11.78 437 E0000285 375 bZIP 1 27 91 SO.9 11.78 437 E0000285 375 bZIP 2 27 81 40.8 833 92 EOOOO368 459 bZIP 1 21 85 51.2 833 92 EOOOO368 459 bZIP 2 21 75 38.5 834 93 E0000369 460 Pkinase 17 269 375.3 -110 834 93 E0000369 460 Pkinase Tyr 17 267 78.6 2O 834 93 E0000369 460 UBA 291 330 23.4 834 93 E0000369 460 KA1 459 507 89.8 835 94 E0000370 461 Pkinase 260 298.7 1340 599 EOOOO667 770 Acyltransferase 174 337 27.2 1179 438 EOOOO668 771 Acyltransferase 2O7 367 67.4 836 95 EOOOO780 853 SRF-TF 59 19.7 836 95 EOOOO780 853 K-box 73 47.6 837 96 EOOOO782. 855 SRF-T 59 13.7 837 96 E0000782. 855 74 51 838 97 EOOOO783 856 R 59 2O2 838 97 EOOOO783 856 74 45.6 1341 600 EOOOO784 857 O3 47.6 839 98 EOOOO785 858 59 19.7 839 98 EOOOO785 858 73 47.6 1294 553 EOOOO788 861 R 59 13.7 1294 553 EOOOO788 861 74 51 1295 554 EOOOO789 862 R 59 13.7 118O 439 E0000790 863 O3 50.5 840 99 E0000791 864 R 59 21 840 99 EOOOO791 864 73 50.5 841 1OO E0000792 865 59 21 842 101 E0000794 867 59 2O2 842 101 E0000794 867 74 45.6 1296 555 EOOOO795 868 R 59 2O2 843 102 EOOOO821. 896 53 32 843 102 EOOOO821. 896 190 319 45.4 1342 6O1 EOOOO880 963 193 262 642 1342 6O1 EOOOO880 963 276 388 36.7 1342 6O1 EOOOO880 963 397 465 944 1342 6O1 EOOOO880 963 HATPase c 510 629 143.2 1297 556 E0000881 964 HAMP 2OO 269 65.7 1297 556 E0000881 964 PAS 283 413 29.7 1297 556 E0000881 964 HisKA 422 490 95.9 1297 556 E0000881 964 HATPase c 535 655 138.8 1343 602 EOOOO883 966 Response reg 276 397 106.2 844 103 E0001051 1141 S4 76 25.1 844 103 E0001051 1141 130 152 19.2 845 104 EOOO1053. 4292 24 155 282.4 846 105 EOOO 1054 1144 133 220 13.4 847 106 EOOO1055 1145 140 285 37.6 847 106 EOOO1055 1145 316 447 48 US 2008/0148432 A1 Jun. 19, 2008

TABLE 11-continued

PEP SEQ NUC ID SEQ NO ID NO GENEID Pfam domain name begin stop score E-value 849 08 PHE0001057 1147 Gamma-thionin 32 8O 77.6 3.SOE-2O 344 603 PHEOOO1063. 1153 CBS 35 175 29.2 1.3OE-OS 344 603 PHEOOO1063. 1153 CBS 193 318 80.8 3.8OE-21 850 09 PHEOOO1064. 1154 AMPKBI 3O4 416 2O8.7 1.2OE-59 345 604 PHEOOO1065. 1155 AMPKBI 298. 412 210.9 2.5OE-60 804 63 PHEOOO1066 1156 AMPKBI 734 852 209.3 7.7OE-60 346 605 PHEOOO1069 1159 bZIP 2 78 35 40.6 4.90E-09 346 605 PHEOOO1069 1159 bZIP 1 82 42 47.6 3.7OE-11 851 10 PHEOOO1071. 1161 AP2 68 31 1492 9.5OE-42 852 11 PHE0001073 1163 Myb DNA-binding 14 61 44.4 3.3OE-10 852 11 PHE0001073 1163 Myb DNA-binding 67 12 SS.S 1.6OE-13 181 440 PHEOOO1074 1164 NAM 19 46 2S5.1 1.3OE-73 853 12 PHE0001075 1165 Myb DNA-binding 22 69 47.5 4.10E-11 853 12 PHE0001075 1165 Myb DNA-binding 75 2O 51 3. SOE-12 347 606 PHEOOO1076 1166 NAM 9 38 293.9 2.7OE-85 854 13 PHE0001077 1167 Myb DNA-binding 227 278 443 3.6OE-10 182 441 PHEOOO1078. 1168 NAM 8 37 3.04.2 2.1OE-88 183 442 PHEOOO1079 1169 NAM 14 42 309 7.6OE-90 348 607 PHEOOO1080 1170 AP2 135 98 137.8 2. SOE-38 184 443 PHEOOO1082 1172 NAM 13 43 299 7.8OE-87 855 14 PHEOOO1083. 1173 AP2 23 86 147.7 2.7OE-41 18S 444 PHEOOO1085. 1175 HLH 381 430 SO.8 4.1OE-12 349 608 PHE0001087. 1177 Homeobox 73 27 74.6 2.90E-19 349 608 PHEOOO1087 1177 HALZ 128 72 813 2.7OE-21 742 1 PHEOOO1089 1179 NAM 14 40 289.7 4.90E-84 350 609 PHEOOO1094. 1184 Zf C2H2 106 28 19.6 O.OO97 856 15 PHEOOO1095. 1185 bZIP 23 87 36.4 8.90E-08 856 15 PHEOOO1095. 1185 bZIP 2 23 77 28.9 16OE-05 857 16 PHEOOO1096 1186 MFMR 1 201 374.1 190E-109 857 16 PHEOOO1096 1186 bZIP 3O3 367 91.7 2.OOE-24 857 16 PHEOOO1096 1186 bZIP 2 3O3 357 31 3.8OE-06 858 17 PHEOOO1097 1187 bZIP 42 95 22.7 O.OOO44 858 17 PHEOOO1097 1187 bZIP 2 42 92 19.3 859 18 PHEOOO1098 1188 GATA 160 195 65.5 6 860 19 PHEOOO1099 1189 MFMR 1 182 333.5 97 860 19 PHEOOO1099 1189 bZIP 257 321 95.2 25 860 19 PHEOOO1099 1189 bZIP 2 257 311 38.9 O8 1351 610 PHEOOO1100 1190 bZIP 187 245 46 O 1351 610 PHEOOO1100 1190 bZIP 2 188 241 44.6 O 861 20 PHEOOO1101 1191 GATA 181 216 65.5 6 862 21 PHE0001107 1197 Myb DNA-binding 24 69 53.4 3 863 22 PHE0001108 1198 Aminotran 1, 2 69 470 61.3 5 864 23 PHE0001109 1199 Aminotran 1, 2 84 469 51.8 2 865 24 PHE0001110 1200 Aminotran 1, 2 61 462 47.4 1 1352 611 PHEOOO1112 1202 Aminotran 1, 2 98 491 46.2 1 1186 445 PHE0001113 1203 Aminotran 5 2O 363 28.9 9 866 25 PHE0001115 1205 Aminotran 1, 2 92 459 285.3 2 866 25 PHE0001115 1205 Cys Met Meta PP 104 339 -275.1 867 26 PHEOOO1120 1210 Histone 62. 137 48.9 867 26 PHEOOO1120 1210 CBFD NFYB HMF 68 132 88.1 868 27 PHEOOO1123. 1213 Histone 92 167 SO.1 868 27 PHEOOO1123. 1213 CBFD NFYB HMF 98 162 89.4 869 28 PHEOOO1125 1215 Histone 94 169 51.5 869 28 PHEOOO1125 1215 CBFD NFYB HMF 1OO 164 92.7 870 29 PHE0001 126 1216 Myb DNA-binding 123 170 44.8 871 30 PHEOOO1127 1217 BURP 299 S19 131.7 872 31 PHEOOO1128 1218 Homeobox 48 103 67.3 872 31 PHEOOO1128 1218. HALZ 104 148 4.1.8 873 32 PHEOOO1130 1220 IlvN 50 233 112.8 873 32 PHEOOO1130 1220 IlvO 237 522 375.3 1353 612 PHEOOO1131. 1221. IlvN 74 244 1892 1353 612 PHEOOO1131. 1221. IlvC 248 395 236.5 874 33 PHE0001132 1222 Aminotran 1, 2 33 401 489.3 743 2 PHEOOO1133 1223 Aminotran 1, 2 44 419 339.8 744 3 PHE0001134 1224 Aminotran 1 2 44 441 408.3 745 4 PHE0001135 1225 Aminotran 1, 2 26 392 492.5 1187 446 PHE0001136. 1226 Aminotran 1, 2 31 399 484.3 875 134 PHE0001138 1228 SapB 1 59 98 21.7 875 134 PHE0001138 1228 SapB 2 1OO 133 23 875 134 PHE0001138 1228 SapB 1 145 183 27.3 875 134 PHE0001138 1228 SapB 2 186 220 29.4 US 2008/0148432 A1 Jun. 19, 2008 34

TABLE 11-continued

PEP SEQ NUC ID SEQ NO ID NO GENEID Pfam domain name begin stop SCO 1188 447 PHEOOO 42 232 Methyltransf 3 66 278 415.7 5. 1354 613 PHEOOO 43 233 HMA 10 70 43.3 7. 876 35 PHEOOO 45 235 F-box 51 O4 33.3 7. 877 36 PHEOOO 46 236 Pkinase 67 321 269.3 6. 878 37 PHEOOO 47 237 UF641 22 59 217.4 2. 879 38 PHEOOO 48 238 yb DNA-binding 221 272 48 2. 1355 614 PHEOOO 49 239 2D 2 87 225.8 8. 880 39 PHEOOO 51 241 RP 355 575 127.6 3. 881 40 PHEOOO 52 242 Ole 58 34 151.1 2. 882 41 PHEOOO 53 243 t l8. Se 34 329.8 4. 883 42 PHEOOO 54 244 S O le 55 104.9 2. 885 44 PHEOOO 57 247 6 592.5 3. 886 45 PHEOOO 58 248 243 179.6 6. 887 46 PEHEOOO 59 249 l8. J 67 103.9 4. 888 47 PHEOOO 67 257 11 211.9 1. 888 47 PHEOOO 67 257 264 64 4. 1356 615 PHEOOO 69 259 OO 108 73.2 7. 889 48 PHEOOO 71 261 OO 62 79.8 7. 890 49 PHEOOO 72 262 OO 111 73.7 5. 1357 616 PHEOOO 76 266 eme Oxygenase 448.6 7. 1358 617 PHEOOO 77 267 eme Oxygenase 384.9 1. 891 SO PHEOOO 79 269 UX IAA 10 299.1 7. 746 5 PHEOOO 81 271 UX IAA 10 296.6 4. 1359 618 PHEOOO 84 274 UX IAA 50 324.3 1. 892 S1 PHEOOO 98 288 209 151.2 2. 892 S1 PHEOOO 98 288 381 155.7 1. 893 52 PHEOOO 99 289 97 144.9 1. 893 52 PHEOOO1199 289 272 151.5 2. 894 53 PHEOOO1208 298 119 147.3 3. 895 54 PHEOOO1211 301 81 117.4 3. 895 54 PHEOOO1211 301 209 110.8 3. 896 55 PHEOOO1212 3O2 17 65.9 1. 896 55 PHEOOO1212 3O2 18 36.6 7. 896 55 PHEOOO1212 3O2 Pkinase Tyr 415 109.7 7. 896 55 PHEOOO1212 3O2 kinase 415 292.2 8. 897 56 PHEOOO1214 304 kinase 400 3O3.4 3. 897 56 PHEOOO1214 304 kinase Tyr 400 1474 3. 899 58 PHEOOO1220 310 kinase 40 3104 2. 900 59 PHEOOO1221 311 kinase 63 307.5 2. 901 60 PHEOOO1225 315 atalase 18 937.9 3. 1298 557 PHEOOO1226 316 atalase 18 937.7 4. 747 6 PHEOOO1227 317 atalase 27 595.4 4. 748 7 PHEOOO1228 3.18 336 103.4 6. 748 7 PHEOOO1228 3.18 kinase 62O 372 8. 748 7 PHEOOO1228 3.18 kinase Tyr 62O 109 1. 902 161 PHEOOO1238 328 Metallothio 2 138.7 1. 903 162 PHEOOO1239 329 Pkinase 309.7 4. 11.90 449 PHEOOO1240 330 H PPase 1072.7 904 163 PHEOOO1241 331 YTH 205.8 9. 905 164 PHEOOO1242 332 hioredoxin 1745 2. 905 164 PHEOOO1242 332 hioredoxin 187 4. 905 164 PHEOOO1242 332 Rp29 172 1. 1.191 450 PHEOOO1243 333 -box 36.2 1. 1.191 450 PHEOOO1243 333 ub 1821 1. 906 165 PHEOOO1244 334 LIM 544 3. 906 165 PHEOOO1244 334 LIM 66 907 166 PHEOOO1245 335 W2 66.1 908 167 PHEOOO1246 337 Zf C3HC4 43.2 749 8 PHEOOO1247 338 Cyclin N 145.4 749 8 PHEOOO1247 338 Cyclin C 52 1360 619 PHEOOO1256 347 Trypsin 18.9 1193 452. PHEOOO1257 348 Fer4 30.6 1193 452. PHEOOO1257 348 Fer4 38.5 910 169 PHEOOO1258 349 Fer4 28.5 910 169 PHEOOO1258 349 Fer4 38.5 911 170 PHEOOO1259 350 Fer4 28.6 911 170 PHEOOO1259 350 Fer4 38.5 1362 621 PHEOOO1265 356 efhand 17.9 11.9S 454 PHEOOO1266 357 efhand 30.3 1363 622 PHEOOO1268 359 DUF788 339.1 1364 623 PHEOOO1270 361 ERO1 748.4 US 2008/0148432 A1 Jun. 19, 2008 35

TABLE 11-continued

PEP SEQ NUC ID SEQ NO ID NO GENEID Pfam domain name begin stop SCO 752 11 EOOO 275 336 Zf C3HC4 257 297 34.6 912 71 EOOO 279 369 Cystatin 41 129 107.9 913 72 EOOO 280 370 Cystatin 49 137 8O.S E-E-E- 914 73 EOOO 282 372 Chal sti Synt C 342 486 13.2 1196 455 EOOO 283 373 Arf6ap 4 120 173.7 915 74 EOOO 284 374 Arf6ap 4 120 175.1 916 75 EOOO 285 375 Arf6ap 6 122 1845 1197 456 EOOO 286 376 IPK 17 274 413.1 -121 917 76 EOOO 291 381 Aa trans 27 433 115.1 917 76 EOOO 291 381 Trp Tyr perm 27 368 -211.7 918 77 EOOO 292 382 Aa trans 40 446 166 918 77 EOOO 292 382 Trp Tyr perm 40 443 -201.1 919 78 EOOO 297 387 DNA photolyase 15 188 237.1 919 78 EOOO 297 387 FAD binding 7 219 497 SO2.9 -148 920 79 EOOO 299 389 DNA photolyase 5 176 224 920 79 EOOO 299 389 FAD binding 7 212 490 464.5 -136 1198 457 EOOO 301 391 Ras 30 192 290 -84 921 8O EOOO 303 393 PRA1 50 2O3 227.8 -65 11.99 458 EOOO 304 394 PRA1 47 197 221 -63 922 81 EOOO 306 396 PRA1 26 177 1715 -48 1365 624 EOOO 311 401 NUDDX 24 160 78.9 -20 923 82 EOOO 312 402 NUDDX 22 153 62.1 -15 924 EOOO 313 403 NUDDX 19 16S 65 -16 925 EOOO 314 404 NUDDX 19 159 71.7 -18 1200 EOOO 316 406 NUDDX 2O 150 69.9 -18 12O1 EOOO 317 407 NUDDX 19 16S 64.4 -16 926 EOOO 332 423 Pkinase 17 272 336.5 -98 926 EOOO 332 423 NAF 330 389 64.3 -16 927 EOOO 333 424 Pkinase 19 274 341.8 -99 927 EOOO 333 424 Pkinase Tyr 19 274 70.1 -18 927 EOOO 333 424 NAF 309 369 108.9 -29 928 EOOO 334 425 Pkinase 14 266 382.4 -112 928 EOOO 334 425 Pkinase Tyr 14 264 86.3 928 EOOO 334 425 UBA 288 327 22.3 928 EOOO 334 425 KA 456 SO4 101S 22 73 1300 EOOO 335 426 Pkinase 46 3OO 359.6 -105 1300 EOOO 335 426 NAF 383 440 103 929 EOOO 337 428 AMPKBI 190 277 111.5 - s 930 EOOO 339 430 AMPKBI 194 281 1231 931 EOOO 343 434 Pkinase 3 268 3416 931 EOOO 343 434 NAF 307 367 124.7 932 EOOO 346 437 Pkinase 2 266 351.8 O2 932 EOOO 346 437 Pkinase Tyr 2 264 72 932 EOOO 346 437 NAF 313 374 99.9 933 EOOO 347 438 Pkinase 5 269 345.1 933 EOOO 347 438 Pkinase Tyr 5 267 80.1 933 EOOO 347 438 NAF 319 378 108.3 934 EOOO 349 440 Pkinase 3 268 341 934 EOOO 349 440 Pkinase Tyr 3 266 74 934 EOOO 349 440 NAF 3O8 369 112.8 935 EOOO 350 441 Pkinase 3 268 3348 935 EOOO 350 441 NAF 307 367 132.7 936 EOOO 354 445 Pkinase 21 276 331.7 936 EOOO 354 445 Pkinase Tyr 21 276 65.1 936 EOOO 354 445 NAF 310 370 94.3 937 EOOO 355 446 Pkinase 7 272 325.1 937 EOOO 355 446 NAF 3OO 368 57.7 938 EOOO 356 447 Pkinase 22 283 325.8 938 EOOO 356 447 NAF 326 385 100.6 939 EOOO 357 448 Pkinase 6 269 3.02.2 939 EOOO 357 448 NAF 309 370 98.4 940 EOOO 358 449 Pkinase 4 266 380.4 940 EOOO 358 449 Pkinase Tyr 4 264 84.4 940 EOOO 358 449 UBA 288 327 22.8 O.OO11 940 EOOO 358 449 KA 454 502 94.9 2.2OE-25 941 EOOO 359 450 Pkinase Tyr 7 267 72.7 1.OOE-18 941 EOOO 359 450 Pkinase 7 269 366.1 S.OOE-107 941 EOOO 359 450 UBA 291 330 20.9 O.OO42 941 EOOO 359 450 KA 460 SO8 84.8 2.4OE-22 942 EOOO 360 451 CAF1 5 248 399.9 3.3OE-117 943 EOOO 361 452 AP2 21 84 135.2 1.6OE-37 US 2008/0148432 A1 Jun. 19, 2008

TABLE 11-continued

PEP SEQ NUC ID SEQ NO ID NO GENEID Pfam domain name begin stop score E-value 944 203 PHEOOO1362. 1453 GATA 220 255 68.2 2.3OE-17 945 204 PHEOOO1363. 1455 NAM 11 135 260.6 2.8OE-75 946 205 PHEOOO1364. 1456 CBS 48 262 43.1 8.40E-10 946 205 PHEOOO1364. 1456 CBS 28S 422 99.1 12OE-26 947 206 PHEOOO1375. 1467 WD40 159 196 50.2 6.3OE-12 947 206 PHEOOO1375. 1467 WD40 201 238 34.4 3.4OE-07 947 206 PHEOOO1375. 1467 WD40 243 28O 41.8 2.1OE-09 947 206 PHEOOO1375. 1467 WD40 28S 322 34.1 4.3OE-07 947 206 PHEOOO1375. 1467 WD40 327 363 15.4 O.19 947 206 PHEOOO1375. 1467 WD40 369 405 10.4 1.4 947 206 PHEOOO1375. 1467 WD40 419 4S5 15.4 0.18 948 207 PHEOOO1377 1469 Pkinase 91 407 238.S 1.3OE-68 949 208 PHEOOO1380 1472 HEAT 82 117 18.3 0.025 949 208 PHEOOO1380 1472 HEAT 158 194 23.5 O.OOO69 949 208 PHEOOO1380 1472 HEAT 197 233 18.6 0.02 949 208 PHEOOO1380 1472 HEAT 236 272 32.2 1.6OE-06 949 208 PHEOOO1380 1472 HEAT 275 311 29.8 8.5OE-06 949 208 PHEOOO1380 1472 HEAT 314 350 26.6 8.OOE-05 949 208 PHEOOO1380 1472 HEAT 353 389 28.8 1.7OE-OS 949 208 PHEOOO1380 1472 HEAT 392 428 26.4 9.2OE-05 949 208 PHEOOO1380 1472 HEAT 431 467 2O O.OO74 949 208 PHEOOO1380 1472 HEAT 47 O SO6 18 O.O3 949 208 PHEOOO1380 1472 HEAT 509 545 21.7 O.OO23 949 208 PHEOOO1380 1472 HEAT 548 S84 22.1 O.OO18 1301 560 PHEOOO1381. 1473 SRF-TF 9 59 99.2 1.1OE-26 1301 560 PHEOOO1381. 1473 K-box 75 172 79.2 1.2OE-2O 754 3 PHEOOO1382. 1474 GRAS 162 SO2 266.8 3.8OE-77 950 209 PHEOOO1385.1477 Cation efflux 1 415 239.1 8.3OE-69 951 210 PHE0001386 1478 Peptidase M20 107 424 213.2 5.1OE-61 951 210 PHEOOO1386. 1478 M20 dimer 214 318 30.6 4.90E-06 952 211 PHE0001389 1481 CN hydrolase 42 219 182.1 12OE-51 953 212 PHE0001392 1484 Pkinase Tyr 89 373 133.9 3.8OE-37 953 212 PHE0001392. 1484 Pkinase 96 373 170.8 3.1OE-48 954 213 PHEOOO1394. 1486 SRF-TF 9 59 116.9 5.1OE-32

954 213 PHE0001394. 1486 K-box 71 174 89 1.3OE-23 955 214 PHEOOO1397 1489 Ran BP1 37 158 159 1.1OE 956 215 PHEOOO1398. 1490 Ran BP1 39 160 185.6 1.1OE 957 216 PHEOOO1405 1497 Nifl N 33 160 273.5 3.8OE 958 217 PHEOOO1411 1503 PSI PsaF 49 227 452.8 3.90E 959 218 PHEOOO1416. 1508 PA 70 171 52.7 1.1OE 959 218 PHE0001416 1508 Peptidase A22B 238 S24 429.6 3.8OE 960 219 PHEOOO1417 1509 NAC 35 92 94.3 3.2OE 962 221 PHE0001421, 1513 ELFV dehydrog N 31 161 2012 2.1OE 962 221 PHE0001421, 1513 ELFV dehydrog 176 4.08 391.9 8.4OE-115 963 222 PHEOOO1426 1518 GATA 91 126 65.1 2.OOE-16 964 223 PHE0001436 1528 Meth synt 1 2 319 594.S 8.7OE-176 964 223 PHE0001436 1528 Meth synt 2 433 757 7S3.8 9.SOE-224 966 225 PHE00014.42 1534 Ubie methyltran 40 256 -109 O.OOO17 966 225 PHEOOO14.42 1534. CMAS 42 295 - 155.1 O.OOO12 966 225 PHE00014.42 1534 Methyltransf 11 1OS 203 105.4 15OE-28 966 225 PHE00014.42 1534 Methyltransf 12 105 201 49.4 1.10E-11 968 227 PHEOOO1452. 1544 ADH N 3O 111 82.8 9.3OE-22 968 227 PHEOOO1452. 1544 ADH Zinc N 142 283 134 3.7OE-37 969 228 PHE0001463. 1555 Homeobox 18 74 86.2 9.2OE-23 969 228 PHEOOO1463. 1555 START 232 456 28O 4.2OE-81 805 64 PHE0001471. 1563 Ammonium transp 36 460 578.1 7.8OE-171 970 229 PHE0001477 1569 Myb DNA-binding 15 62 49.1 1.3OE-11 970 229 PHE0001477 1569 Myb DNA-binding 68 113 50.6 4.7OE-12 971 230 PHEOOO1481 1573 GAF 195 339 35.3 1.90E-07 971 230 PHEOOO1481 1573 HisKA 37S 440 36.4 8.90E-08 971 230 PHE0001481 1573 Response reg 641 762 14.8 2.8OE-05 972 231 PHE0001483. 1575 Glyco hydro 16 29 210 416.9 2.6OE-122 972 231 PHEOOO1483. 1575 XET C 234 284 99.8 7.3OE-27 973 232 PHEOOO1516 1607 SelR 87 209 274.8 1. SOE-79 1367 626 PHEOOO1519 1610 WD40 9 47 29.3 1.2OE-OS 1367 626 PHEOOO1519 1610 WD40 52 89 34.3 3.7OE-07 1367 626 PHEOOO1519 1610 WD40 94 131 49.1 1.3OE-11 1367 626 PHEOOO1519 1610 WD40 136 173 53 9.OOE-13 1367 626 PHEOOO1519 1610 WD40 178 215 48.8 1.6OE-11 1367 626 PHEOOO1519 1610 WD40 22O 256 8.1 2.7 1302 561 PHE0001521. 1612 Xan ur permease 36 443 195.6 1.OOE-SS

US 2008/0148432 A1 Jun. 19, 2008

TABLE 11-continued

PEP SEQ NUC ID SEQ NO ID NO GENEID Pfam domain name begin stop score E-value 266 525 PHE0003742 4044 NAD binding 4 11 251 -54.1 3.6OE-05 267 526 PHE0003743. 4046 adh short 7 176 -21 9.7OE-OS 267 526 PHEOOO3743. 4046 RmlD Sub bind 8 338 - 131 2.1OE-OS 267 526 PHE0003743. 4046 Epimerase 9 273 2S7.7 2.1OE-74 267 526 PHE0003743 4046 Polysacc synt 2 9 323 -159.7 O.OOO63 267 526 PHEOOO3743. 4046 3Beta HSD 10 297 -57.4 7.8OE-09 267 526 PHE0003743 4046 NAD binding 4 11 248 -56.6 5.2OE-05 476 735 PHEOOO3788 41.22 NAM 2 97 1642 3.1OE-46 334 593 PHEOOO3815 4288 HAMP 2OO 269 65.7 1.3OE-16 334 593 PHEOOO3815 4288 PAS 283 413 29.7 9.1OE-06 334 593 PHEOOO3815 4288. HisKA 422 490 95.9 1.OOE-2S 334 593 PHEOOO3815 4288. HATPase c 535 655 141.9 16OE-39 268 527 PHEOOO38.25 4.192 NAM 14 140 297 3.1OE-86 269 528 PHEOOO3839 4208 Zf LSD1 27 51 45.6 1SOE-10 269 528 PHEOOO3839 4208 Zif LSD1 66 90 54.8 2. SOE-13 269 528 PHEOOO3839 4208 Zif LSD1 104 128 58.4 2.1OE-14 270 529 PHEOOO3885 4266 Globin 14 154 96.2 8.90E-26 271 530 PHEOOO3886. 4267 Globin 13 152 113.7 4.7OE-31 477 736 PHEOOO3899 4284 PHD 18O 227 60.2 6.2OE-15 272 531 PHE0003927 4321 PAD porph 1 4 368 694.5 6.7OE-2O6 803 62 PHEOOO3945. 4522 Zf C3HC4 146 187 39.2 1.2OE-08 478 737 PHEOOO3948 4528 MFS 1 84 S2O 28.5 2.OOE-OS 273 532 PHE0003959 4544 Response reg 79 202 93.9 4.3OE-2S 273 532 PHEOOO3959 4544 CCT 695 736 57.3 4.6OE-14 274 533 PHE0003961. 4662 ketoacyl-synt 47 295 266.4 4.90E-77 274 533 PHE0003961. 4662 Ketoacyl-synt C 3O3 458 209.1 9.4OE-60 275 534 PHEOOO3965. 4553 bZIP 1 277 335 39.8 8.1OE-09 275 534 PHEOOO3965. 4553 bZIP 2 277 334 40.9 3.8OE-09 276 535 PHE0003966 4554 FAE 3-kCoA syn1 2O 365 699 3.1OE-2O7 277 536 PHEOOO3977 4565 HEM4 56 285 85.7 1.3OE-22 479 738 PHEOOO3981. 4568 PK 22 368 809.5 16OE-240 479 738 PHEOOO3981. 4568 PK C 382 S13 221.1 2.2OE-63 28O 539 PHEOOO3993 4579 Na Cal ex 147 28O 119.1 1.1OE-32 338 597 PHEOOO3995 4581 Succ DH flav C 6 136 265.6 8.6OE-77 283 542 PHE0003997 4583 Epimerase 12 226 -40 O.OO 283 542 PHEOOO3997 4583 NmirA 12 309 475.3 6.SOE-140 481 740 PHE0003998. 4584 Auxin inducible 1 86 77.5 3.6OE-2O 287 546 PHEO004.003 4589 Transferase 22 447 2S4.6 1.8OE-73 290 549 PHEO004006 4592 Abhydrolase 1 164 425 36.8 6.6OE-08 482 741 PHEO004008 4594 Trypsin 112 272 16.7 2. SOE-OS 292 551 PHEO004010 4596 Myb DNA-binding 347 392 43.1 8.40E-10 339 598 PHEOOO4022 4657 PHD 2O2 2S2 SS.9 1.2OE-13

TABLE 12 accession gathering Pfam domain name number cutoff domain description 2-Hacid dh PFOO389.18 13.2 D-isomer specific 2-hydroxyacid dehydrogenase, catalytic domain 2-Hacid dh C PFO2826.6 -75.7 D-isomer specific 2-hydroxyacid dehydrogenase, NAD binding domain 3Beta HSD PFO1073.8 -135.9 3-beta hydroxysteroid dehydrogenase isomerase family 3 5 exonuc PFO1612.10 -32 3'-5' exonuclease AAA PFOOOO4.17 10 ATPase family associated with various cellular activities (AAA) AA kinase PFOO696.16 -40 Amino acid kinase family AA permease PFOO324.10 -120.8 Amino acid permease ABC1 PFO3109.6 -27.6 ABC1 family ABC train PFOOOOS.14 8.6 ABC transporter ADH N PFO8240.1 -14.5 Alcohol dehydrogenase GroES-like domain ADH zinc N PFOO107.15 23.8 Zinc-binding dehydrogenase AMP-binding PF005O1.15 O AMP-binding enzyme AMPKBI PFO4739.4 25 5'-AMP-activated protein kinase, beta Subunit, complex-interacting region US 2008/0148432 A1 Jun. 19, 2008 47

TABLE 12-continued

accession gathering Pfam domain name number cutoff domain description AP2 FOO847.9 O AP2 domain APS kinase FO1583.9 25 Adenylylsulphate kinase ARID FO1388.10 -8 ARID/BRIGHT DNA binding domain AT hook FO2178.7 14.2 AT hook motif AUX IAA FO2309.6 -83 AUXIAA family Aa trans FO1490.7 -128.4 Transmembrane amino acid transporter protein Abhydrolase 1 FOOS61.9 5.5 alpha/beta hydrolase fold Acetyltransf 1 FOOS83. 18.6 Acetyltransferase (GNAT) family Acyltransferase FO1553. 6 Acyltransferase Aldedh FOO171. -295 Aldehyde dehydrogenase family Aldo ket red FOO248. -97 Aldo/keto reductase family Alpha-amylase FOO128. -93 Alpha amylase, catalytic domain Alpha adaptinc2 FO2883.9 -12 Adaptin C-terminal domain Aminotran 1, 2 FOO155.9 -57.5 Aminotransferase class I and II Aminotran 3 FOO2O2.10 -2O7.6 Aminotransferase class-III Aminotran 5 FOO266.8 -92.9 Aminotransferase class-V Ammonium transp FOO909.10 -144 Ammonium Transporter Family Ank FOOO23.17 21.6 Ankyrin repeat Annexin FOO1918 Annexin Arf6ap FO1412.8 -17 Putative GTPase activating protein for Arf ASn synthase FOOf33.10 -52.8 Asparagine synthase Asp FOOO26.13 -186.1 Eukaryotic aspartyl protease Auxin inducible FO2S19.4 -15 Auxin responsive protein Auxin resp 25 Auxin response factor B12D 25 B12D protein 26.5 B3 DNA binding domain FO1603.8 -210 Protein phosphatase 2A regulatory B subunit (B56 family) BAH FO1426.6 BAH domain BRO1 FO3097.6 25 BRO1-like domain BURP FO31815 -52 BURP domain Bromodomain FOO439.13 8.9 Bromodomain CAF1 FO4857.8 -100.5 CAF1 family ribonuclease CBFD NFYB HMF FOO8O8.12 18.4 Histone-like transcription factor (CBF/NF Y) and archaeal histone CBS 15.8 CBS domain pair CCT 25 CCT motif CH 22.5 Calponin homology (CH) domain CMAS FO2353.9 -177.9 Cyclopropane-fatty-acyl-phospholipid syntinase CN hydrolase FOO795.11 -13.9 Carbon-nitrogen hydrolase CTP synth N FO6418.2 25 CTP synthase N-terminus CTP transf 2 FO1467.15 -11.8 Cytidylyltransferase Carb kinase FO1256.7 -66.3 Carbohydrate kinase Catalase FOO199.8 -229 Catalase Cation efflux FO1545.10 -95.7 Cation efflux family Chal sti Synt C FO2797.5 -6.1 Chalcone and stilbene synthases, C terminal domain Chromo FOO385.11 27.5 chromo (CHRromatin Organisation MOdifier) domain Citrate synt FOO285.10 -101.5 Citrate synthase CobW C FO7683.3 18 Cobalamin synthesis protein cobW C terminal domain ComA FO2679.5 25 (2R)-phospho-3-sulfolactate synthase (ComA) CorA FO1544.8 -61.3 CorA-like Mg2+ transporter protein Cpn10 FOO166.11 -7.8 Chaperonin 10 Kod subunit Cpnó0 TCP1 FOO118.13 -223.4 TCP-1/cpnó0 chaperonin family Cu-Oxidase FOO394.11 -18.9 Multicopper oxidase Cu-Oxidase 2 FO7731.3 -5.8 Multicopper oxidase Cu-Oxidase 3 FO7732.4 10 Multicopper oxidase Cyclin C FO2984.7 -13 Cyclin, C-terminal domain Cyclin N FOO13412 -14.7 Cyclin, N-terminal domain Cyclotide FO3784.3 25 Cyclotide family Cys Met Meta PP FO1 OS3.9 -278.4 Cys/Met metabolism PLP-dependent enzyme Cystatin FOOO31.10 17.5 CyStatin domain DAO FO1266.11 -36.5 FAD dependent oxidoreductase DNA photolyase FOO875.7 -10 DNA photolyase DSPC FOOf82.9 -21.8 Dual specificity phosphatase, catalytic domain US 2008/0148432 A1 Jun. 19, 2008 48

TABLE 12-continued

accession gathering Pfam domain name number cutoff domain description DUF125 FO1988.8 -10.1 integral membrane protein DUF125 DUF1423 FO7227.1 25 Protein of unknown function (DUF1423) DUF1530 FO7O60.1 25 ProFAR isomerase associated DUF1685 FO7939.1 25 Protein of unknown function (DUF1685) DUF246 FO3138.4 -15 Plant protein family DUF250 FO3151.6 125 Domain of unknown function, DUF250 DUF296 FO3479.4 -11 Domain of unknown function (DUF296) DUF393 FO41342 25 Protein of unknown function, DUF393 DUF581 FO4570.4 -3.1 Protein of unknown function (DUF581) DUF6 FOO892.9 30 integral membrane protein DUF6 DUF641 FO48S9.2 25 Plant protein of unknown function (DUF641) DUF760 FOSS42.1 25 Protein of unknown function (DUF760) DUF788 FOS620.1 25 Protein of unknown function (DUF788) Dehydrin FOO257.8 -4.4 Dehydrin Di19 FO5605.2 25 Drought induced 19 protein (Di19) Dirigent FO3O18.4 25 Dirigent-like protein DnaJ FOO226.18 -8 DnaJ domain E1 dh FOO676.9 -90 Dehydrogenase E1 component E2F TDP FO2319.9 17 E2F/DP family winged-helix DNA binding domain EB FO3271.6 25 EB1-like C-terminal motif EF1 GNE FOOf3 6.8 2O EF-1 guanine nucleotide exchange domain ELFV dehydrog FOO2O8.10 -27 Glutamate? Leucine/Phenylalanine? Valine dehydrogenase ELFV dehydrog N C FO2812.7 31.8 Glu/Leu/Phe(Valdehydrogenase, dimerisation domain ERO1 F04137.5 -179.5 Endoplasmic Reticulum Oxidoreductin 1 (ERO1) ER29 FO7749.2 1O.S Endoplasmic reticulum protein ERp29, C erminal domain Epimerase FO1370.10 -46.3 NAD dependent epimerase? dehydratase amily F-box FOO646.20 12.4 F-box domain FAD binding 3 FO1494.8 -136.6 FAD binding domain FAD binding 4 -8.1 FAD binding domain FAD binding 7 25 FAD binding domain of DNA photolyase FAE 3-kCoA syn1 25 Fatty acid elongase 3-ketoacyl-CoA synthase 1 FA desaturase FOO487.13 -46 Fatty acid desaturase FBA 1 FO7734.2 -39.4 F-box associated FBPase FOO316.9 -170.3 Fructose-1-6-bisphosphatase FGGY N FOO370.10 -104.7 FGGY family of carbohydrate kinases, N erminal domain FHA FOO498. 25 FHA domain Fera. FOOO37. 4Fe-4S binding domain GAF FO1590. 23 GAF domain GAT FO3127.4 GAT domain GATA FOO32O. 28.5 GATA zinc finger GATase FOO117. -38.1 utamine amidotransferase class-I GATase 2 FOO310. -106.2 utamine amidotransferases class-II GFO IDH MocA FO1408. -7.2 xidoreductase family, NAD-binding OSSmann fol GFO IDH MocA C FO2894.7 xidoreductase family, C-terminal 8. lphabeta domain GH3 FO3321.3 -3.36 H3 auxin-responsive promoter GIDA FO1134.11 -226.7 lucose inhibited division protein A GRAS FO3S14.4 -78 RAS family transcription factor GRIM-19 FO621.2.1 25 RIM-19 protein GSHPx FOO255.9 -16 utathione peroxidase GST C FOOO43.13 22.3 utathione S-transferase, C-terminal O main GST N C FO2798.8 14.6 utathione S-transferase, N-terminal O main GTP EFTU FOOOO9.14 ongation factor Tu GTP binding domain GTP EFTU D2 FO314413 25 ongation factor Tu domain 2 GTP EFTU D3 FO3143.6 14.3 ongation factor Tu C-terminal domain Gamma-thionin FOO3O4.10 9.6 amma-thionin family Gln-synt C FOO120.13 -124 utamine synthetase, catalytic domain Gln-synt N FO3951.8 utamine synthetase, beta-Grasp domain Globin FOOO42.11 -8.8 obin Glyco hydro 1 FOO232.8 -301.8 ycosyl hydrolase family 1 Glyco hydro 14 FO1373.7 -231.4 ycosyl hydrolase family 14 US 2008/0148432 A1 Jun. 19, 2008 49

TABLE 12-continued

accession gathering Pfam domain name number cutoff domain description yco hydro 16 FOO722.9 -65 Glycosyl family 16 yco hydro 38 C FO1074.11 -125.3 Glycosyl hydrolases family 38 N-terminal domain yco hydro 38C FO7748.2 -93.1 Glycosyl hydrolases family 38 C-terminal domain yco transf 20 FOO982.9 -243.6 Glycosyltransferase family 20 ycogen Syn FOS693.2 -492.3 Glycogen synthase ycolytic FOO274.8 -158 Fructose-bisphosphate aldolase class-I ycos transf 1 FOOS34.9 -7.3 Glycosyltransferases group 1 ycos transf 2 FOOS35.14 17.6 Glycosyltransferase family 2 yoxalase FOO903.14 12.1 Glyoxalase, Bleomycin resistance protein Dioxygenase Superfamily Ot 1 FO4178.2 25 Got1-like family p dh C FO2800.8 -64.1 Glyceraldehyde 3-phosphate dehydrogenase, C-terminal domain p dh N FOOO44.11 -74.2 Glyceraldehyde 3-phosphate dehydrogenase, NAD binding domain HALZ FO2183.7 Homeobox associated leucine Zipper HAMP FOO672.13 HAMP domain HATPase c FO2S18.13 istidine kinase-, DNA gyrase B-, and SP90-like ATPase HEAT FO2985.9 HEAT repeat HEM4 FO2602.5 Uroporphyrinogen-III synthase HemD HGTP anticodon FO3129.9 Anticodon binding domain HIO933 like FO3486.4 IO933-like protein HLH FOOO10.15 elix-loop-helix DNA-binding domain HMA FOO403.14 eavy-metal-associated domain HMG box FOO505.8 MG (high mobility group) box HSF DNA-bind FOO447.7 SF-type DNA-binding HSP20 FOOO11.9 sp20/alpha crystallin family H PPase norganic H-- pyrophosphatase tlene OXygenase deme oxygenase Hexapep Bacterial transferase hexapeptide (three repeats) Hexokinase 1 Hexokinase Hexokinase 2 Hexokinase HisKA FOOS12.13 is Kinase A (phosphoacceptor) domain Hist deacetyl FOO850.9 Histone deacetylase domain Histone FOO125. Core histone H2AH2B, H3/H4 Homeobox FOOO46. Homeobox domain FO1627. Hpt domain FOO702. haloacid dehalogenase-like hydrolase FOO463.9 Socitrate lyase family FO1652.8 Eukaryotic initiation factor 4E FO3770.6 nositol polyphosphate kinase FO1450.8 -33.8 Acetohydroxy acid isomeroreductase, catalytic domain wN FO7991. -75.8 Acetohydroxy acid isomeroreductase, catalytic domain inhibitor I29 FO8246. 4.9 Cathepsin propeptide inhibitor domain (I29) On trans FOOS2O.18 -4.5 on transport protein Soamylase N FO2922.7 -6.5 Soamylase N-terminal domain acalin FO1419.6 2O acalin-like lectin domain FO2373.11 mC domain FO2375.6 25 imN domain K-box FO1486.7 K-box region KA1 FO2149.9 25 Kinase associated domain 1 KH 1 FOOO13.17 8.1 KH domain Kelch 1 FO1344.13 Kelch motif Kelch 2 FO7646.4 2O Kelch motif Ketoacyl-synt C FO28O1.10 -54-9 Beta-ketoacylsynthase, C-terminal domain Kunitz legume FOO197.8 -32 Trypsin and protease inhibitor LEA 5 FOO477.7 25 Small hydrophilic plant seed protein LIM FOO412.10 LIM domain LRR 2 FO7723.2 8.7 Leucine Rich Repeat Lactamase B FOO753.15 22.3 Metallo-beta-lactamase Superfamily Ldh 1 C FO2866.6 -13 actate/malate dehydrogenase, alphabeta C-terminal domain Ldh 1 N FOOOS 6.11 -31.3 actate/malate dehydrogenase, NAD binding domain US 2008/0148432 A1 Jun. 19, 2008 50

TABLE 12-continued

accession gathering Pfam domain name number cutoff domain description Lectin legA FOO138.7 19 Legume lectins alpha domain Lectin legB FOO139.9 -77 Legume lectins beta domain Lig chan FOOO60.1 6 8.2 Ligand-gated ion channel Lipase GDSL FOO657.1 1 10.9 GDSL-like Lipase? Acylhydrolase M20 dimer FO7687.3 12 Peptidase dimerisation domain MAP1 LC3 FO2991.5 -18.8 Microtubule associated protein 1A, 1B, light chain 3 MFMR FO7777.1 -46.7 G-box binding protein MFMR MFS 1 FO7690.4 23.5 Major Facilitator Superfamily MIP FOO230.8 -62 Major intrinsic protein Malic M FO3949.4 -143.9 Malic enzyme, NAD binding domain Mat FO1554.8 59.6 Mat Metallophos FOO149.1 6 22 Calcineurin-like phosphoesterase Metallothio 2 FO1439.7 -3 Metallothionein Meth synt 1 FO8267.1 -167.8 Cobalamin-independent synthase, N terminal domain Meth synt 2 FO1717.7 -15S Cobalamin-independent synthase, Catalytic domain Methyltransf 11 FO8241.1 17.1 Methyltransferase domain Methyltransf 12 FO8242.1 21.4 Methyltransferase domain Methyltransf 2 FOO891.7 -103.8 O-methyltransferase Methyltransf 3 FO1596.7 -120.6 O-methyltransferase Mpv17 PMP22 FO4117.2 -5.4 Mpv17/PMP22 family MtN3 silv FO3O83.5 -0.8 MtN3/saliva family Myb DNA-binding FOO249.1 8 19.1 Myb-like DNA-binding domain NAC FO1849.6 NAC domain NAD binding 4 FO7993.1 -87.7 Malesterility protein NAF FO3822.4 25 NAF domain NAM FO2365.5 -19 No apical meristem (NAM) protein NDK FOO334.8 -59.9 Nucleoside diphosphate kinase NIF FO3031.7 -81 NL1 interacting factor-like phosphatase NPH3 FO3OOO.4 25 NPH3 family NTF2 FO2136. Nuclear transport factor 2 (NTF2) domain NTP transferase FOO483. -90.5 Nucleotidyl transferase NUDIX FOO293. NUDIX domain Na Cal eX FO1699. 25 Sodium calcium exchanger protein Nifl N FO1592.6 -13 Nifl-like N terminal domain NmirA FOS368.2 -90.6 Nmir A-like family Orn Arg deC N FO2784.6 -76 Pyridoxal-dependent decarboxylase, pyridoxal binding domain Orn DAP Arg deC FOO278. -34.9 Pyridoxal-dependent decarboxylase, C erminal sheet domain Oxidored FMN FOOf24.8 -147.7 NADH:flavin oxidoreductase,NADH oxidase family PA FO2225. O 13 PA domain PAD porph FO4371.4 -180.8 Porphyromonas-type peptidyl-arginine deiminase PARP FOO644.9 -55.5 Poly(ADP-ribose) polymerase catalytic domain PAS FOO989. 2O PAS fold PB1 FOOS64. 12.1 PB1 domain BD FOOf86. 12.1 P21-Rho-binding domain CI FO1399. 25 PCI domain DZ FOO595. 12.1 PDZ domain (Also known as DHR or GLGF) EP-utilizers FOO391. 2 10 PEP-utilising enzyme, mobile domain EP-utilizers C FO2896.7 -173 PEP-utilising enzyme, TIM barrel domain EPcase FOO311.7 25 Phosphoenolpyruvate carboxylase GAM FOO3OO. 1 -3 Phosphoglycerate mutase family HD FOO628. 6 25.9 PHD-finger K FOO224. O -244 Pyruvate kinase, barrel domain K C FO2887.5 -44 Pyruvate kinase, alphabeta domain MSR FO1625.9 -62 Peptide methionine sulfoxide reductase P2C FOO481. O -44 Protein phosphatase 2C PDK N FO1326.8 -87 Pyruvate phosphate dikinase, PEP pyruvate binding domain RA1 FO32O8.8 25 PRA1 family protein SI PsaF FO2507.5 25 Photosystem I reaction centre subunit III TR2 FOO854.1 1 -50 POT family UA FO1472.8 2.2 PUA domain Peptidase A22B FO4258.3 -137.3 Signal peptide peptidase Peptidase C1 FOO112.1 1 -115.8 Papain family cysteine protease Peptidase C15 FO1470.7 -100 Pyroglutamyl peptidase US 2008/0148432 A1 Jun. 19, 2008 51

TABLE 12-continued

accession gathering Pfam domain name number cutoff domain description Peptidase M20 C FO1546.16 -14.4 Peptidase family M20/M25/M40 Peptidase S10 C FOO4SO.11 -198 Serine carboxypeptidase Peptidase S41 C FO3572.7 -25.8 Peptidase family S41 PfkB C FOO294.12 -67.8 pfkB family carbohydrate kinase Phytochrome C FOO360.9 11 Phytochrome region Pkinase C FOOO69.14 -70.8 Protein kinase domain Pkinase C C FOO433.11 14 Protein kinase C terminal domain Pkinase Tyr C FO7714.4 65 Protein tyrosine kinase Polysacc synt 2 C FO2719.5 -176 Polysaccharide biosynthesis protein Pro CA C FOO484.8 -45 Carbonic anhydrase Pro dh C FO1619.7 -120.5 Proline dehydrogenase Pyr redox C FOOO70.16 5 Pyridine nucleotide-disulphide oxidoreductase Pyr redox 2 FO7992.2 -20 Pyridine nucleotide-disulphide oxidoreductase Pyr redox dim FO2852.11 -13 Pyridine nucleotide-disulphide oxidoreductase, dimerisation domain Pyridoxal deC FOO282.8 -158.6 Pyridoxal-dependent decarboxylase conserved domain RHD3 FO5879.2 25 Root hair defective 3 GTP-binding protein (RHD3) RIO1 FO1163.11 -89.1 RIO1 family RRM 1 FOOO76.10 15.2 RNA recognition motif. (a.k.a. RRM, RBD, or RNP domain) RTC FO1137.11 -36.9 RNA 3'-terminal phosphate cyclase RTC insert FOS189.3 25 RNA 3'-terminal phosphate cyclase (RTC), insert domain RWP-RK FO2O42.5 25 RWP-RK domain Ran BP1 FOO638.8 -38 RanBP1 domain Ras FOOO71.11 18 Ras family Remorin C FO3763.3 25 Remorin, C-terminal region Response reg FOOO72.11 -14.4 Response regulator receiver domain Reticulon FO2453.7 -40 Reticulon Ribonuclease T2 FOO445.8 -53 Ribonuclease T2 family Ribosomal L1 FOO687.10 -101 Ribosomal protein LlpfL10e family Ribosomal L10e FOO826.7 25 Ribosomal L10 Ribosomal L12 FOOS42.8 25 Ribosomal protein L7/L12 C-terminal domain Ribosomal L19e FO1280.9 -28 Ribosomal protein L19e Ribosomal L39 FOO832.9 25 Ribosomal L39 protein Ribosomal L7Ae FO1248.13 Ribosomal protein L7 Ae/L30e/S12e/Gadd45 family Ribosomal S11 FOO411.7 Ribosomal protein S11 Ribosomal S17 FOO366.9 1.7 Ribosomal protein S17 Ribosomal S2 FOO318.9 -22 Ribosomal protein S2 Ribosomal S27 FO1599.8 50 Ribosomal protein S27a Rieske FOO355.15 Rieske [2Fe-2S domain RmlD Sub bind FO4321.6 -1718 RmlD binding domain RuBisCO small FOO101.9 -20.1 Ribulose bisphosphate carboxylase, Small chain Rubrerythrin FO2915.7 -4.8 Rubrerythrin SAM 1 FOO536.17 11.3 SAM domain (Sterile alpha motif) SAM 2 FO7647.5 2O SAM domain (Sterile alpha motif) SPC25 FO67.03.1 25 Microsomal signal peptidase 25kDa. subunit (SPC25) SPX C FO3105.9 -20 SPX domain SRF-TF C FOO319.8 11 SRF-type transcription factor (DNA binding and dimerisation domain) START FO1852.8 25 START domain SapB 1 FOS1844 2O Saposin-like type B, region 1 SapB 2 FO3489.5 2O Saposin-like type B, region 2 SecY FOO344.9 -210 eubacterial secY protein SelR FO1641.8 -66.5 SelR domain Sigma/O r1 2 FOO140.9 25 Sigma-70 factor, region 1.2 Sigma70 r2 FO4S42.3 11 Sigma-70 region 2 Sigma70 r3 FO4539.4 10 Sigma-70 region 3 Sigma70 ra. FO4S4S.S 20.7 Sigma-70, region 4 Sina FO3145.6 -48.4 Seven in absentia protein family Steroid dh FO2S44.6 -44.7 3-oxo-5-alpha-steroid 4-dehydrogenase Suc Fer-like FO6999.2 -42.4 Sucrase? ferredoxin-like Succ DH flav C FO2910.9 Fumarate reductase? succinate dehydrogenase flavoprotein C-terminal domain US 2008/0148432 A1 Jun. 19, 2008 52

TABLE 12-continued

accession gathering Pfam domain name number cutoff domain description Sucrose synth FOO862.9 -134 Sucrose synthase Sugar tr FOOO83.12 -85 Sugar (and other) transporter Synaptobrevin FOO957.9 25 Synaptobrevin TPP enzyme C FO2775.9 19.7 Thiamine pyrophosphate enzyme, C terminal TPP binding domain TPP enzyme M FOO2O5.11 -23.9 Thiamine pyrophosphate enzyme, central omain TPP enzyme N FO2776.7 -70 Thiamine pyrophosphate enzyme, N terminal TPP binding domain Thiolase C FO28O3.6 -30.7 Thiolase, C-terminal domain Thiolase N FOO108.11 -129.5 Thiolase, N-terminal domain Thioredoxin FOOO85.8 -25.7 Thioredoxin Tic22 FO4278.2 25 Tic22-like family Transaldolase FOO923.8 -49 Transaldolase Transferase FO2458.5 -161.2 Transferase family Transket pyr FO2779.12 -50 Transketolase, pyridine binding domain Transketolase C FO2780.9 -15.5 Transketolase, C-terminal domain Transketolase N FOO456.10 -98 Transketolase, thiamine diphosphate binding domain Trehalase FO1204.8 25 Trehalase Trehalase Ca-bi FO7492.1 2O Neutral trehalase Ca2+ binding domain Trehalose PPase FO2358.6 -49.4 Trehalose-phosphatase Trp Tyr perm FO3222.3 -232.6 Tryptophantyrosine permease family Trp syntA FOO290.10 -149.8 Tryptophan synthase alpha chain Trypsin FOOO89.13 -33.2 Trypsin Tub FO1167.7 -98 Tub family Tubulin FOOO91.14 -55.7 Tubulin/FtsZ family, GTPase domain Tubulin C FO3953.6 -10 Tubulin/FtsZ family, C-terminal domain UBA FOO627.18 2O.S UBATS-N domain UDPGP FO1704.7 -265.2 UTP-glucose-1-phosphate uridylyltransferase UDPGT -151 UDP-glucoronosyl and UDP-glucosyl transferase UPF0057 FO1679.7 25 Uncharacterized protein family UPF0057 UbiA FO1040.8 UbiA prenyltransferase family Ubie methyltran FO1209.8 -117 ubiECOQ5 methyltransferase family Usp FOO582.15 25.7 Universal stress protein family VHS FOOfSO.8 -13.2 VHS domain FO5678.3 25 VQ motif FO2O2O.7 25 eIF4-gammatelF5/eIF2-epsilon WD40 FOO400.19 21.4 WD domain, G-beta repeat WHEP-TRS FOO458.9 10 WHEP-TRS domain WRKY FO3106.5 25 WRKY DNA-binding domain Wzy C FO4932.4 25 O-Antigen Polymerase XET C FO6955.2 11.4 Xyloglucan endo-transglycosylase (XET) C-terminus Xan ur permease FOO860.10 -151.2 Permease family YL1 FOS764.3 25 YL1 nuclear protein FO8265.1 18.6 YL1 nuclear protein C-terminal domain YTH FO4146.5 25 YT521-B-like family Yippee FO3226.4 25 Yippee putative zinc-binding protein YeF N FO3853.3 25 YeF-related protein N-terminus ZF-HD dimer FO4770.2 25 ZF-HD protein dimerisation region Zip FO2535.10 -28 ZIP Zinc transporter adh short FOO106.13 -46.6 short chain dehydrogenase bZIP 1 FOO170.10 16.5 bZIP transcription factor bZIP 2 FO7716.4 15 Basic region leucine Zipper cNMP binding FOOO27.17 20.6 Cyclic nucleotide-binding domain cobW FO2492.8 -10 CobWHypB/UreG, nucleotide-binding domain efhand FOOO36.19 17.5 EF hand ketoacyl-synt FOO109.14 -73.6 Beta-ketoacylsynthase, N-terminal domain malic FOO390.8 25 Malic enzyme, N-terminal domain p450 FOOO67.11 -105 Cytochrome P450 peroxidase FOO141.12 -10 Peroxidase tRNA-synt 2b FOOS87.14 -40.5 RNA synthetase class II core domain (G, H, P, S and T) ubiquitin FOO240.12 19.4 Ubiquitin family Zf-B box FOO643.13 11.1 B-box zinc finger US 2008/0148432 A1 Jun. 19, 2008

TABLE 12-continued accession gathering Pfam domain name number cutoff domain description Zf C2H2 PFOOO96.14 19 Zinc finger, C2H2 type Zf C3HC4 PFOOO97.12 16.9 Zinc finger, C3HC4 type (RING finger) Zf-DOf PFO27015 25 Dof domain, Zinc finger Zf-LSD1 PFO6943.2 25 LSD1 Zinc finger

EXAMPLE 8 in Example 4. Transgenic plant cells of corn, soybean, cotton, canola, wheat and rice are transformed with recombinant Selection of Transgenic Plants with Enhanced Agro DNA for expressing each of the homologs identified in nomic Trait(s) Example 4. Plants are regenerated from the transformed plant 0126. This example illustrates the preparation and identi cells and used to produce progeny plants and seed that are fication by selection of transgenic seeds and plants derived screened for enhanced water use efficiency, enhanced cold from transgenic plant cells of this invention where the plants tolerance, increased yield, enhanced nitrogen use efficiency, and seed are identified by screening a having an enhanced enhanced seed protein and enhanced seed oil. Plants are iden agronomic trait imparted by expression of a protein selected tified exhibiting enhanced traits imparted by expression of the from the group including the homologous proteins identified homologous proteins.

SEQUENCE LISTING The patent application contains a lengthy “Sequence Listing section. A copy of the “Sequence Listing is available in electronic form from the USPTO web site (http://seqdata.uspto.gov/?pageRequest=docDetail&DocID=US20080148432A1). An electronic copy of the “Sequence Listing will also be available from the USPTO upon request and payment of the fee set forth in 37 CFR1.19(b)(3).

1-22. (canceled) acid sequence in the group consisting of SEQID NO: 1138, 23. A plant cell with stably integrated, recombinant DNA 1139, 1321, 1429, 1430, 1431 and 1432. comprising a promoter that is functional in plant cells and that 26. A plant cell of claim 23 further comprising DNA is operably linked to DNA encoding a seven-in-absentia pro expressing a protein that provides tolerance from exposure to tein, an herbicide applied at levels that are lethal to a wild type of said plant cell. wherein said plant cell is present in a plant or seed that 27. A plant cell of claim 26 wherein the agent of said (a) exhibits an enhanced trait as compared to control herbicide is a glyphosate, dicamba, or glufosinate compound. plants that do not have said recombinant DNA; and 28. A transgenic plant comprising a plurality of the plant (b) is derived from a progenitor plant or seed that was cell of claim 23. selected as having said enhanced trait from a popula 29. A transgenic seed comprising a plurality of the plant tion of plants or seeds that have said recombinant cell of claim 23. DNA, wherein said enhanced trait is selected from 30. A transgenic seed of claim 29 from a corn, soybean, group consisting of enhanced water use efficiency, cotton, canola, alfalfa, wheat or rice plant. enhanced cold tolerance, increased yield, enhanced 31. A transgenic pollen grain comprising a haploid deriva nitrogen use efficiency, enhanced seed protein and tive of a plant cell of claim 23. enhanced seed oil, and 32. A method for manufacturing non-natural, transgenic wherein a portion of said population exhibits said seed that can be used to produce a crop of transgenic plants enhanced trait. with an enhanced trait resulting from expression of stably 24. The plant cell of claim 23 wherein said DNA encoding integrated, recombinant DNA comprising a promoter that is a seven-in-absentia protein is from a plant, bacteria or yeast (a) functional in plant cells and (b) is operably linked to DNA and encodes a seven-in-abstentia protein having at least one from a plant, bacteria or yeast that encodes a seven-in-absen domain of amino acids in a sequence that exceeds the Pfam tia protein; wherein said enhanced trait is selected from the gathering cutoff for amino acid sequence alignment with group of enhanced traits consisting of enhanced water use protein domain family identified by the Pfam name Sina; efficiency, enhanced cold tolerance, increased yield, wherein the Pfam gathering cutoff for the Sina protein enhanced nitrogen use efficiency, enhanced seed protein and domain family is 48.4. enhanced seed oil; wherein said method comprises: 25. A plant cell of claim 23 wherein said protein has an (a) screening a population of plants for said enhanced trait amino acid sequence with at least 90% identity to an amino and said recombinant DNA, wherein individual plants in US 2008/0148432 A1 Jun. 19, 2008 54

said population can exhibit said trait at a level less than, (c) verifying that said recombinant DNA is stably inte essentially the same as or greater than the level that said grated in said selected plants, trait is exhibited in control plants which do not express (d) analyzing tissue of a. selected plant to determine the the recombinant DNA production of a seven-in-absentia protein in plant cells of claim 23, and (b) selecting from said population one or more plants that (e) collecting seed from a selected plant. exhibit the trait at a level greater than the level that said trait is exhibited in control plants, ck