US00961553OB2

(12) United States Patent (10) Patent No.: US 9,615,530 B2 May (45) Date of Patent: Apr. 11, 2017

(54) COTTON VARIETY 14R938B2XF 7,855,326 B2 12/2010 Feng et al. 7,858,764 B1 12/2010 Huberet al. (71) Applicant: MONSANTO TECHNOLOGY LLC, 7,939,721 B2 5/2011 Arnevik et al. St. Louis, MO (US) 8,071,735 B2 12/2011 Cerny et al. s 8,119,380 B2 2/2012 Weeks et al. 8.420,888 B2 4/2013 Feng et al. (72) Inventor: O. Lloyd May, Tifton, GA (US) 8,435,743 B2 5/2013 Cerny et al. 8,629,323 B2 1/2014 Weeks et al. (73) Assignee: Monsanto Technology LLC, St. Louis, 8,735,661 B2 5 2014 Brinker et al. MO (US) 9,024,115 B2 5, 2015 Brinker et al. 9,133,473 B2 9/2015 Huber et al. 9,179,620 B2 11/2015 Ma (*) Notice: Subject to any disclaimer, the term of this 2012/03 11730 A1* 12/2012 NE ...... A23D 9.00 patent is extended or adjusted under 35 800,260 U.S.C. 154(b) by 0 days. FOREIGN PATENT DOCUMENTS (21) Appl. No.: 14/968,531 WO WO 2004/072235 8, 2004 (22) Filed: Dec. 14, 2015 OTHER PUBLICATIONS (65) Prior Publication Data U.S. Appl. No. 14/968,639, filed Dec. 14, 2015, McGowen. US 2016/0174499 A1 Jun. 23, 2016 U.S. Appl. No. 14/968,547, filed Dec. 14, 2015, May. U.S. Appl. No. 14/996,093, filed Jan. 14, 2016, McGowen. Related U.S. ApplicationO O Data ReleasedBowman Betweenet al., “Pedigrees 19702005, of UplandMississippi and AgriculturalPima Cotton & Cultivars Forestry (60) Provisional application No. 62/095,531, filed on Dec. Experiment Station, Bulletin. 1155. Dec. 2006, 22, 2014 Eshed et al., “Less-than-addictive epistatic interactions of quanti s tative trait loci in tomato,' Genetics, 143: 1807-1817, 1996. Fehr (ed), "Backcross Method.” In: Principles of Cultivar Devel (51) Int. Cl. opment, vol. 1: Theory and Technique, pp. 360-376, 1987. AOIH IM00 (2006.01) Steet.” Theor:i. Appl.E.g. Genet. seqiya,ingerprinting :323-32O, in Sugar AOIH 5/00 (2006.01) Poehlman (ed), “Breeding Corn (Maize).” In: Breeding Field Crops, AOIH 5/10 (2006.01) 3rd Ed., AVI Publishing Company, Westport, Connecticut, pp. CI2N 5/82 (2006.01) 469-481, 1987. (52) U.S. Cl. Poehlman (eds), "Backcross Breeding.” In: Breeding Field Crops, 4th Ed., pp. 172-175, 1995. CPC ------r r A0IH 5/10 (2013.01) Rieger et al., In: Glossary of Genetics and Cytogenetics, Classical (58) Field of Classification Search and Molecular, Springer-Verlag, Berlin, p. 116, 1976. None Sprague et al. (Eds.), “8-1.1.2 Backcrossing.” In: Corn and See application file for complete search history. Improvement, 3' Ed., Madison, WI, pp. 472-473, 1998. Marchosky et al., “Bollgard(R) and Bollgard IIR Efficacy in Near (56) References Cited Isogenic Lines of DP50. Upland Cotton in Arizona.” Arizona Cotton Report, The University of Arizona College of Agriculture U.S. PATENT DOCUMENTS and Life Sciences, 2001. Phipps et al., “Performance of Roundup FlexC) cotton varieties in 4.940,835. A 7, 1990 Shah the North Mississippipp River Delta,, pp.pp 874-880, 2006 Beltwide 5.338,544 A 8, 1994 Donovan Cotton Conferences, San Antonio, Texas, Jan. 3-6, 2006. 5.45 i. 514 A 9, 1995 Boudet et al. Virginia Cotton Production Guide 2011, College of Agriculture and 5,523.520 A 6/1996 Hunsperger et al. Life Sciences, Virginia Tech, 2011. 5,633,435 A 5/1997 Barry Variety specific information as indicated in transmittal letter of Mar. 5,717,084 A 2f1998 Herrera-Estrella et al. 3, 2016. Information Disclosure Statement of U.S. Appl. No. 5,728,925 A 3, 1998 Herrera-Estrella et al. 14/968,531. 5,850,019 A 12/1998 Maiti et al. 5,981,834 A 11/1999 John et al. * cited by examiner 6,051,753. A 4/2000 Comai et al. g f 3.3. R et al. Primary Examiner — Mykola V. Kovalenko 648954 B 12/2002 Ea. (74) Attorney, Agent, or Firm — Dentons US LLP 6,660,911 B2 12/2003 Fincher et al. 6,740,488 B2 5/2004 Rangwala et al. (57) ABSTRACT 6943.2826.893.826 BiB1 9,5, 2005 HillvardNE et al. The invention relates to the novel cotton variety designated 6949,696 B2 9, 2005 Fincher et al. 14R938B2XF. Provided by the invention are the seeds, 7,022,896 B1 4/2006 Weeks et al. plants, plant parts and derivatives of the cotton variety 7,064.249 B2 6/2006 Corbinet al. 14R938B2XF. Also provided by the invention are methods 7,105,729 B2 9/2006 Mitchell et al. of using cotton variety 14R938B2XF and products derived 7,112,665 B1 9/2006 Leemans et al. therefrom. Still further provided by the invention are meth 7,112,725 B2 9, 2006 Fincher et al. ds f duci 1 b h 7,141,722 B2 11/2006 Fincher et al. O s or produc1ng cotton p ants by crossing the cotton 7,223,907 B2 5, 2007 Huber et al. variety 14R938B2XF with itself or another cotton variety 7,381,861 B2 6/2008 Cerny et al. and plants and seeds produced by Such methods. 7,770,830 B1 8/2010 Langenecker et al. 7,812,224 B2 10/2010 Weeks et al. 22 Claims, No Drawings US 9,615,530 B2 1. 2 COTTON VARIETY 14R938B2XF as one parent, the second generation (F) hybrid cotton plant grown from the seed of the F hybrid plant, and the seeds of This application claims the benefit of U.S. provisional the F hybrid plant. application No. 62/095,531, filed Dec. 22, 2014, which is In a further aspect of the invention, a composition is herein incorporated by reference in its entirety. 5 provided comprising a seed of cotton variety 14R938B2XF comprised in plant seed growth media. In certain embodi BACKGROUND OF THE INVENTION ments, the plant seed growth media is a soil or synthetic cultivation medium. In specific embodiments, the growth 1. Field of the Invention medium may be comprised in a container or may, for The present invention relates generally to the field of 10 example, be soil in a field. Plant seed growth media are well known to those of skill in the art and include, but are in no cotton breeding. In particular, the invention relates to the way limited to, Soil or synthetic cultivation medium. Advan novel cotton variety 14R938B2XF. tageously, plant seed growth media can provide adequate 2. Description of Related Art physical Support for seeds and can retain moisture and/or The goal of a commercial cotton breeding program is to 15 nutritional components. Examples of characteristics for soils develop new, unique and Superior cotton varieties. In cotton, that may be desirable in certain embodiments can be found, important traits include higher fiber (lint) yield, earlier for instance, in U.S. Pat. Nos. 3,932,166 and 4,707,176. maturity, improved fiber quality, resistance to diseases and Synthetic plant cultivation media are also well known in the insects, tolerance to drought and heat, and improved agro art and may, in certain embodiments, comprise polymers or nomic traits. The breeder initially selects and crosses two or hydrogels. Examples of such compositions are described, for more parental lines, followed by generation advancement example, in U.S. Pat. No. 4.241,537. Still yet another aspect of the invention is a method of and selection, thus producing many new genetic combina producing cotton seeds comprising crossing a plant of the tions. The breeder can theoretically generate billions of cotton variety 14R938B2XF to any second cotton plant, different genetic combinations via this procedure. including itself or another plant of the variety 14R938B2XF. 25 In particular embodiments of the invention, the method of SUMMARY OF THE INVENTION crossing comprises the steps of a) planting seeds of the cotton variety 14R938B2XF; b) cultivating cotton plants One aspect of the present invention relates to seed of the resulting from said seeds until said plants bear flowers; c) cotton variety 14R938B2XF. The invention also relates to allowing fertilization of the flowers of said plants; and, d) plants produced by growing the seed of the cotton variety 30 harvesting seeds produced from said plants. 14R938B2XF, as well as the derivatives of such plants. As Still yet another aspect of the invention is a method of used herein, the term “plant' includes plant cells, plant producing hybrid cotton seeds comprising crossing the cot ton variety 14R938B2XF to a second, distinct cotton plant protoplasts, plant cells of a tissue culture from which cotton which is nonisogenic to the cotton variety 14R938B2XF. In plants can be regenerated, plant calli, plant clumps, and plant particular embodiments of the invention, the crossing com cells that are intact in plants or parts of plants. Such as pollen, 35 prises the steps of a) planting seeds of cotton variety flowers, seeds, bolls, leaves, stems, and the like. 14R938B2XF and a second, distinct cotton plant, b) culti Another aspect of the invention relates to a tissue culture Vating the cotton plants grown from the seeds until the plants of regenerable cells of the cotton variety 14R938B2XF, as bear flowers; c) cross pollinating a flower on one of the two well as plants regenerated therefrom, wherein the regener plants with the pollen of the other plant, and d) harvesting ated cotton plant expresses all the physiological and mor 40 the seeds resulting from the cross pollinating. Still yet another aspect of the invention is a method for phological characteristics of a plant grown from the cotton developing a cotton plant in a cotton breeding program seed designated 14R938B2XF. comprising: obtaining a cotton plant, or its parts, of the Yet another aspect of the current invention is a cotton variety 14R938B2XF; and b) employing said plant or parts plant of the cotton variety 14R938B2XF comprising at least 45 as a source of breeding material using plant breeding tech a first transgene, wherein the cotton plant is otherwise niques. In the method, the plant breeding techniques may be capable of expressing all the physiological and morphologi selected from the group consisting of recurrent selection, cal characteristics of the cotton variety 14R938B2XF. In mass selection, bulk selection, backcrossing, pedigree particular embodiments of the invention, a plant is provided breeding, genetic marker-assisted selection and genetic that comprises a single locus conversion. A single locus 50 transformation. In certain embodiments of the invention, the conversion may comprise a transgenic gene which has been cotton plant of variety 14R938B2XF is used as the male or introduced by genetic transformation into the cotton variety female parent. 14R938B2XF or a progenitor thereof. A transgenic or non Still yet another aspect of the invention is a method of transgenic single locus conversion can also be introduced by producing a cotton plant derived from the cotton variety 14R938B2XF, the method comprising the steps of: (a) backcrossing, as is well known in the art. In certain embodi 55 preparing a progeny plant derived from cotton variety ments of the invention, the single locus conversion may 14R938B2XF by crossing a plant of the cotton variety comprise a dominant or recessive allele. The locus conver 14R938B2XF with a second cotton plant; and (b) crossing sion may confer potentially any desired trait upon the plant the progeny plant with itself or a second plant to produce a as described herein. progeny plant of a Subsequent generation which is derived Still yet another aspect of the invention relates to a first 60 from a plant of the cotton variety 14R938B2XF. In one generation (F) hybrid cotton seed produced by crossing a embodiment of the invention, the method further comprises: plant of the cotton variety 14R938B2XF to a second cotton (c) crossing the progeny plant of a Subsequent generation plant. Also included in the invention are the F hybrid cotton with itself or a second plant; and (d) repeating steps (b) and plants grown from the hybrid seed produced by crossing the (c) for at least 2-10 additional generations to produce an cotton variety 14R938B2XF to a second cotton plant. Still 65 inbred cotton plant derived from the cotton variety further included in the invention are the seeds of an F. 14R938B2XF. Also provided by the invention is a plant hybrid plant produced with the cotton variety 14R938B2XF produced by this and the other methods of the invention. US 9,615,530 B2 3 4 Plant variety 14R938B2XF-derived plants produced by this be reproduced by planting and growing seeds of the variety and the other methods of the invention described herein may, under self-pollinating or sib-pollinating conditions, as is in certain embodiments of the invention, be further defined known to those of skill in the agricultural arts. Variety as comprising the traits of plant variety 14R938B2XF given 14R938B2XF shows no variants other than what would in Table 1. 5 normally be expected due to environment or that would The use of the word “a” or “an' when used in conjunction occur for almost any characteristic during the course of with the term “comprising in the claims and/or the speci repeated sexual reproduction. The results of an objective fication may mean “one.” but it is also consistent with the description of the variety are presented below, in Table 1. meaning of “one or more,” “at least one.” and “one or more Those of skill in the art will recognize that these are typical values that may vary due to environment and that other than one.” 10 Other objects, features and advantages of the present values that are Substantially equivalent are within the scope invention will become apparent from the following detailed of the invention. description. It should be understood, however, that the detailed description and the specific examples, while indi TABLE 1. cating specific embodiments of the invention, are given by 15 Phenotypic Description of Variety 14R938B2XF way of illustration only, since various changes and modifi cations within the spirit and scope of the invention will Species: Gossypium hirsuttin L. become apparent to those skilled in the art from this detailed Areas of Adaptation: description. Eastern Delta DETAILED DESCRIPTION OF THE Central Plains INVENTION General: The invention provides, in one aspect, methods and Plant Habit: Intermediate composition relating to plants, seeds and derivatives of the Foliage: Intermediate 25 Stem Lodging: Intermediate cotton variety 14R938B2XF. Cotton variety 14R938B2XF Fruiting Branch: Normal was developed from an initial cross of 08Y619B2R2*4/ Growth: Intermediate DP0912-BRD-BC1-F2-0132. The breeding history of the Leaf Color: Medium Green variety can be Summarized as follows:

Generation Year Description Cross 2010 Cross was made in Leland, MS between donor parent DP0912 BRD-BC1-F2-0132 (containing the gene event MON 88701 for dicamba and glufosinate tolerance) and recurrent parent 08Y619B2R2 (containing the gene events MON 15985 for epidopteron tolerance and MON 88913 for glyphosate tolerance). Plants were grown in Leland, MS and advanced using bulk based on event of interest selection. BC 2010 Cross was made in Leland, MS using an F. plant selection and 08Y619B2R2 as the recurrent parent. BCF 2010 Plants were grown in Leland, MS and advanced using plant selection for event of interest. BC, 2010 Cross was ma e in Leland, MS using a BCIF plant selection and 08Y619B2R2 as the recurrent parent. BC2F 2011 Plants were grown in Juana Diaz, PR and a vanced using plant Selection base on event of interest and MABC recovery of the recurrent parent. BC 2011 Cross was ma e in Juana Diaz, PR using a BC2F1 plant selection and 08Y619B2R2 as he recurrent parent. BCF 2011 Plants were grown in uana Diaz, PR and a vanced using plant Selection base on event of interest selection and MABC recovery of the recurren parent. BCF2 2012 Plants were gro wn in Juana Diaz, PR and a vanced using plant Selection base on homozygosity for the intended events. BCF 2012 Plants were grown in uana Diaz, PR and a vanced using bulk based on gene purity.

Advanced Testing 55 TABLE 1-continued Phenotypic Description of Variety 14R938B2XF Generation Year Selection Boll Shape: Length is more than width BCF 2013 selected based on the lint yield, lint percent, fiber Boll Breadth: Broadest at middle quality and trueness to type with the recurrent 60 Maturity: parent. BCFs 2014 selected based on the lint yield, lint percent, fiber Days till maturity: 137 quality and trueness to type with the recurrent Plant: parent. Cm to 1. Fruiting Branch (from cotyledonary node): 33.0 65 No. of Nodes to 1. Fruiting Branch (excluding cotyledonary node): 6.1 The cotton variety 14R938B2XF has been judged to be Mature Plant Height (from cotyledonary node to terminal): 134 uniform for breeding purposes and testing. The variety can US 9,615,530 B2 5 6 TABLE 1-continued TABLE 1-continued Phenotypic Description of Variety 14R938B2XF Phenotypic Description of Variety 14R938B2XF Leaf (Upper most, fully expanded leaf): Nematodes, Insects and Pests: Bollworm: Resistant Type: Normal Pink bollworm: Resistant Pubescence: Sparse Tobacco bud worm: Resistant Nectaries: Present Stem: These are typical values, Values may vary due to environment. Other values that are 10 substantially equivalent are within the scope of the invention. Stem Pubescence: Intermediate Cotton variety 14R938B2XF is Bollgard IIR), Roundup Glands: Ready(R) Flex, and dicamba and glufosinate tolerant cotton variety (containing the gene events MON 15985, MON Leaf: Normal 88913, and MON 88701). MON 15985 produces the Stem: Normal 15 Cry1Ac and Cry2Ab proteins which give protection from Calyx lobe (normal is absent): Normal feeding damage caused by lepidopteran insect pests. Cry1Ac Flower: and Cry2Ab came from the bacteria Bacillus thuringiensis (subsp. Kurstaki). MON 88913 produces a 5-enolpyruvyl Petals: Cream shikimate-3-phosphate synthase protein from Agrobacte Pollen: Cream rium sp. strain CP4 (CP4 EPSPS), which confers tolerance Petal Spot: Absent to glyphosate, the active ingredient in the Roundup(R) family Seed: of agricultural herbicides. MON 88701 produces a dicamba monooxygenase from Stenotrophomonas maltophilia and a Seed Index (g/100 seeds, fuzzy basis): 9.2 phosphinothricin acetyltransferase from Streptomyces Lint Index (g lint 100 seeds): 6.7 hygroscopicus, which confers dicamba and glufosinate tol Boll: 25 erance. Events MON 15985, MON 88913, and MON 88701 are the subjects of U.S. Pat. Nos. 7,223,907, 7,381,861, and Lint percent, picked: 41.7 8.735,661, respectively, the disclosures of which are incor Number of seeds per boll: 34 porated herein by reference. Events MON 15985, MON Grams Seed Cotton per Boll: 5.5 88913, and MON 88701 are also covered by one or more of Boll Type: open 30 the following patents: U.S. Pat. Nos. 5,717,084; 5,728,925; Fiber Properties: 5,850,019; 6,051,753: 6,083,878; 6,489,542: 6,660,911; 6,949,696; 6,943,282; 7,022,896; 7,064.249; 7,112,665; Method (HVI or other): HVI 7,112,725; 7,141,722; 7,223,907; 7,381,861; 7,700,830; Length (inches, 2.5% SL): 1.14 7,812,224; 7,855,326; 7,858,764; 7,939,721; 8,071,735; Uniformity (%): 84.3 35 8,119,380; 8,420,888; 8,629,323. Strength, T1 (g/tex): 30.3 The performance characteristics of cotton variety Elongation, E1 (%): 11.6 14R938B2XF were also analyzed and comparisons were Micronaire: 4.3 made with competing varieties. The results of the analysis are presented below, in Table 2. TABLE 2 Performance Data for Variety 14R938B2XF Entries Compared YLD BE LP MIC LNTH UNIF STRN EL PHT 14R938B2XF 1,702 43.31 4.84 1.11 84.13 29.12 8.84 38.1 DP 1137 B2RF 1,685 42.44 4.73 1.13 84.46 29.42 8.57 37.5 Deviation 16.19 O.87 0.11 -0.02 -0.32 -0.3 0.27 O.S8 Significance :::::: :::::: :::::: -- -- : # Obs 22 22 12 10 12 12 12 19 Years 1 1 1 1 1 1 1 1 Win Percent 55 86 17 10 17 33 67 32 Test Mean 1,707 41.65 4.65 1.16 84.56 31.4 8.71 37 14R938B2XF 1,472 42.49 4.68 1.1 82.8 29.15 10.74 40.9 DP 1050 B2RF 1,456 41.25 4.54 1.14 83.16 31.15 1O-38 40 Deviation 16.36 1.24 0.14 -0.04 -0.35 -2 O.36 O.96 Significance :::::: :::::: :::::: :::::: :::::: :::::: : # Obs 51 52 37 35 37 37 36 36 Years 2 2 2 2 2 2 2 2 Win Percent 59 85 19 6 25 14 69 50 Test Mean 1,478 41.15 4.57 1.13 83.3 30.7 10.64 39 14R938B2XF 1,481 42.36 4.6 1.1 82.92 29.43 11.1 40.4 14R911B2XF 1,598 41.19 4.54 1.13 83.83 30.13 11.33 36.5 Deviation -117.75 1.17 O.06 -0.03 -0.91 -0.7 -0.23 3.85 Significance :::::: :::::: :::::: :::::: :::::: :::::: :::::: # Obs 60 60 46 44 46 46 46 42 Years 2 2 2 2 2 2 2 2 Win Percent 32 83 37 14 11 16 24 14 Test Mean 1,521 40.95 4.52 1.14 83.43 31.01 11.03 38.6 14R938B2XF 1,481 42.36 4.6 1.1 82.92 29.43 11.1 40.4 14R91SB2XF 1,559 40.92 4.42 1.13 83.57. 30.12 11.29 37.4 Deviation -78.21 1.44 0.18 -0.03 -0.66 -0.69 -0.2 2.98

US 9,615,530 B2 9 10 TABLE 2-continued Performance Data for Variety 14R938B2XF Entries Compared YLD BE LP MIC LNTH UNIF STRN EL PHT Test Mean 1,502 40.95 4.52 1.13 83.39 30.98 10.97 38.5 14R938B2XF 1,486 42.42 4.59 1.1 82.9S 29.38 11.02 40.3 14R914B2XF 1,548 39.75 4.52 1.14 8368. 31.76 11.24 36.6 Deviation -61.61 2.66 O.07 -0.03 -0.72 -2.38 -0.21 3.62 Significance -- :::::: -- :::::: :::::: :::::: k: :::::: # Obs 59 59 45 43 45 45 45 41 Years 2 2 2 2 2 2 2 2 Win Percent 44 97 33 12 14 O 25 7 Test Mean 1,528 40.95 4.51 1.14 83.46. 30.98 10.94 38.5 14R938B2XF 1,507 42.41 4.67 1.1 82.89 29.14 10.7 41 14R949B2XF 1,470 41.06 4.47 1.14 83.37 30.02 1O.S 39.7 Deviation 36.9 1.35 0.2 -0.04 -0.48 -0.88 O2 1.33 Significance :::::: :::::: :::::: :::::: :::::: k: :::::: # Obs S4 55 39 37 39 39 39 39 Years 2 2 2 2 2 2 2 2 Win Percent 59 93 10 8 26 23 81 33 Test Mean 1,522 40.96 4.SS 13 83.4 30.83 10.64 39 14R938B2XF 1,328 41.89 4.54 .1 82.32 29.57 11.93 41.9 PHY 499 WRF 1,402 40.17 4.57 12 83.28 32.69 12.07 41.8 Deviation -74.47 1.72 -0.03 -0.03 -0.96 -3.11 -0.15 O.OS Significance :::::: :::::: :::::: :::::: :::::: # Obs 39 40 35 35 35 35 35 25 Years 1 1 1 1 1 1. 1 Win Percent 38 88 52 9 6 3 35 52 Test Mean 1,396 40.62 4.5 12 82.89 30.86 11.87 39.3 14R938B2XF 1,328 41.89 4.54 .1 82.32 29.57 11.93 41.9 ST 4946GLB2 1,455 39.6 4.53 12 82.99 31.98 11.64 37 Deviation -127.SS 2.3 0.01 -0.03 -0.66 -2.41 O.28 4.81 Significance :::::: :::::: :::::: :::::: :::::: k: :::::: # Obs 39 40 35 35 35 35 35 25 Years 1 1 1 1 1 1. 1 Win Percent 33 88 38 14 17 9 67 4 Test Mean 1,396 40.62 4.5 12 82.89 30.86 11.87 39.3 **, *, + Significant at P × 0.01, 0.05, or 0.10, respectively

35 I. BREEDING COTTON VARIETY 14R938B2XF Cotton plants can be crossed by either natural or mechani cal techniques. Natural pollination occurs in cotton either by One aspect of the current invention concerns methods for self pollination or natural cross pollination, which typically crossing the cotton variety 14R938B2XF with itself or a is aided by pollinating organisms. In either natural or second plant and the seeds and plants produced by Such 40 artificial crosses, flowering and flowering time are important methods. These methods can be used for propagation of the considerations. cotton variety 14R938B2XF or can be used to produce The cotton flower is perfect in that the male and female hybrid cotton seeds and the plants grown therefrom. A structures are in the same flower. The crossed or hybrid seed hybrid plant can be used as a recurrent parent at any given can be produced by manual crosses between selected par stage in a backcrossing protocol during the production of a 45 ents. Floral buds of the parent that is to be the female can be single locus conversion of the cotton variety 14R938B2XF. emasculated prior to the opening of the flower by manual The variety of the present invention is well suited to the removal of the male anthers. At flowering, the pollen from development of new varieties based on the elite nature of the flowers of the parent plants designated as male, can be genetic background of the variety. In selecting a second manually placed on the Stigma of the previous emasculated plant to cross with 14R938B2XF for the purpose of devel 50 flower. Seed developed from the cross is known as first oping novel cotton varieties, it will typically be desired to generation (F) hybrid seed. Planting of this seed produces choose those plants which themselves exhibit one or more F hybrid plants of which half their genetic component is selected desirable characteristics. Examples of potentially from the female parent and half from the male parent. desired characteristics include higher fiber (lint) yield, ear Segregation of genes begins at meiosis thus producing lier maturity, improved fiber quality, resistance to diseases 55 second generation (F) Seed. Assuming multiple genetic and insects, tolerance to drought and heat, and improved differences between the original parents, each F seed has a agronomic traits. unique combination of genes. Any time the cotton variety 14R938B2XF is crossed with Self-pollination occurs naturally in cotton with no another, different, variety, first generation (F) cotton prog manipulation of the flowers. For the crossing of two cotton eny are produced. The hybrid progeny are produced regard 60 plants, it may be beneficial to use artificial hybridization. In less of characteristics of the two varieties produced. As such, artificial hybridization, the flower used as a female in a cross an F1 hybrid cotton plant may be produced by crossing is manually cross pollinated prior to maturation of pollen 14R938B2XF with any second cotton plant. The second from the flower, thereby preventing self fertilization, or cotton plant may be genetically homogeneous (e.g., inbred) alternatively, the male parts of the flower are emasculated or may itself be a hybrid. Therefore, any F hybrid cotton 65 using a technique known in the art. Techniques for emas plant produced by crossing cotton variety 14R938B2XF culating the male parts of a cotton flower include, for with a second cotton plant is a part of the present invention. example, physical removal of the male parts, use of a genetic US 9,615,530 B2 11 12 factor conferring male sterility, and application of a chemi pedigree selection, modified pedigree selection, mass selec cal gametocide to the male parts. tion, recurrent selection and backcrossing. For artificial hybridization employing emasculation, The complexity of inheritance influences choice of the flowers that are expected to open the following day are breeding method. Backcrossbreeding is used to transfer one selected on the female parent. The buds are swollen and the or a few favorable genes for a highly heritable trait into a corolla is just visible through the calyx or has begun to desirable variety. This approach has been used extensively emerge. Usually no more than two buds on a parent plant are for breeding disease-resistant plant varieties. Various recur prepared, and all self-pollinated flowers or immature buds rent selection techniques are used to improve quantitatively are removed with forceps. Special care is required to remove inherited traits controlled by numerous genes. The use of immature buds that are hidden under the stipules at the leaf 10 recurrent selection in self-pollinating crops depends on the axil, and could develop into flowers at a later date. The ease of pollination, the frequency of Successful hybrids from flower is grasped between the thumb and index finger and each pollination, and the number of offspring from each the location of the Stigma determined by examining the Successful cross. sepals. The calyx is removed by grasping a sepal with the Each breeding program should include a periodic, objec forceps, pulling it down and around the flower, and repeating 15 tive evaluation of the efficiency of the breeding procedure. the procedure until the five sepals are removed. The exposed Evaluation criteria vary depending on the goal and objec corolla is removed with care to avoid injuring the Stigma. tives, but should include gain from selection per year based Cross-pollination can then be carried out using, for example, on comparisons to an appropriate standard, overall value of petri dishes or envelopes in which male flowers have been the advanced breeding lines, and number of Successful collected. Desiccators containing calcium chloride crystals varieties produced per unit of input (e.g., per year, per dollar can be used in Some environments to dry male flowers to expended, etc.). obtain adequate pollen shed. Promising advanced breeding lines are thoroughly tested Either with or without emasculation of the female flower, and compared to appropriate standards in environments hand pollination can be carried out by removing the stamens representative of the commercial target area(s) for generally and pistill with a forceps from a flower of the male parent and 25 three or more years. The best lines are candidates for new gently brushing the anthers against the Stigma of the female commercial varieties. Those still deficient in a few traits may flower. Access to the stamens can be achieved by removing be used as parents to produce new populations for further the front sepal and keel petals, or piercing the keel with selection. closed forceps and allowing them to open to push the petals These processes, which lead to the final step of marketing away. Brushing the anthers on the Stigma causes them to 30 and distribution, may take as much as eight to 12 years from rupture, and the highest percentage of Successful crosses is the time the first cross is made. Therefore, development of obtained when pollen is clearly visible on the stigma. Pollen new varieties is a time-consuming process that requires shed can be checked by tapping the anthers before brushing precise forward planning, efficient use of resources, and a the stigma. Several male flowers may have to be used to minimum of changes in direction. obtain suitable pollen shed when conditions are unfavorable, 35 A most difficult task is the identification of individuals or the same male may be used to pollinate several flowers that are genetically Superior, because for most traits the true with good pollen shed. genotypic value is masked by other confounding plant traits Cross-pollination is more common within rows than or environmental factors. One method of identifying a between adjacent rows; therefore, it may be beneficial to superior plant is to observe its performance relative to other grow populations with genetic male sterility on a square grid 40 experimental plants and to one or more widely grown to create rows in all directions. For example, single-plant standard varieties. Single observations are generally incon hills on 50-cm centers may be used, with subdivision of the clusive, while replicated observations provide a better esti area into blocks of an equal number of hills for harvest from mate of genetic worth. bulks of an equal amount of seed from male-sterile plants in The goal of plant breeding is to develop new, unique and each block to enhance random pollination. 45 superior cotton varieties. The breeder initially selects and There are numerous steps in the development of any crosses two or more parental lines, followed by repeated novel, desirable plant germplasm. Plant breeding begins selfing and selection, producing many new genetic combi with the analysis and definition of problems and weaknesses nations. Each year, the plant breeder selects the germplasm of the current germplasm, the establishment of program to advance to the next generation. This germplasm is grown goals, and the definition of specific breeding objectives. The 50 under unique and different geographical, climatic and soil next step is selection of germplasm that possess the traits to conditions, and further selections are then made, during and meet the program goals. The goal is to combine in a single at the end of the growing season. The varieties which are variety an improved combination of desirable traits from the developed are unpredictable. This unpredictability is parental germplasm. These important traits may include because the breeder's selection occurs in unique environ resistance to diseases and insects, tolerance to drought and 55 ments, with no control at the DNA level (using conventional heat, tolerance to herbicides, improvements in fiber traits breeding procedures), and with millions of different possible and numerous other agronomic traits that may be desirable genetic combinations being generated. A breeder of ordinary to the farmer or end user. skill in the art cannot predict the final resulting lines he Choice of breeding or selection methods depends on the develops, except possibly in a very gross and general mode of plant reproduction, the heritability of the trait(s) 60 fashion. The same breeder cannot produce the same variety being improved, and the type of variety used commercially twice by using the exact same original parents and the same (e.g., F hybrid variety, pureline variety, etc.). For highly selection techniques. This unpredictability results in the heritable traits, a choice of superior individual plants evalu expenditure of large amounts of research monies to develop ated at a single location will be effective, whereas for traits Superior new cotton varieties. with low heritability, selection should be based on mean 65 Pureline cultivars, such as generally used in cotton and values obtained from replicated evaluations of families of many other crops, are commonly bred by hybridization of related plants. Popular selection methods commonly include two or more parents followed by selection. The complexity US 9,615,530 B2 13 14 of inheritance, the breeding objectives and the available tives, Wageningen (Ed), Center for Agricultural Publishing resources influence the breeding method. The development and Documentation, 1979: Fehr, In: Principles of variety of new varieties requires development and selection, the development. Theory and Technique (Vol 1) and Crop Spe crossing of varieties and selection of progeny from Superior cies Soybean (Vol 2), Iowa State Univ., Macmillian Pub. COSSS. 5 Co., NY, 360-376, 1987: Fehr, In: Soybeans: Improvement, Pedigree breeding and recurrent selection breeding meth Production and Uses,’ 2d Ed., Monograph 16:249, 1987). ods are used to develop varieties from breeding populations. Additionally, with any of the methods disclosed above, Breeding programs combine desirable traits from two or mutagenesis can be utilized to increase the diversity of the more varieties or various broad-based sources into breeding gene pool that is available in the breeding program. pools from which varieties are developed by selfing and 10 selection of desired phenotypes. The new varieties are Proper testing should detect any major faults and establish evaluated to determine which have commercial potential. the level of superiority or improvement over current vari Pedigree breeding is commonly used for the improvement eties. In addition to showing Superior performance, there of self-pollinating crops. Two parents which possess favor must be a demand for a new variety that is compatible with able, complementary traits are crossed to produce an F. An 15 industry standards or which creates a new market. The F. population is produced by selfing one or several F plants. introduction of a new variety will incur additional costs to Selection of the best individuals may begin in the F. the seed producer, the grower, processor and consumer, for population or later depending upon objectives of the special advertising and marketing, altered seed and com breeder; then, beginning in the F, the best individuals in the mercial production practices, and new product utilization. best families can be selected. Replicated testing of families 20 The testing preceding release of a new variety should take can begin in the F or F generation to improve the effec into consideration research and development costs as well as tiveness of selection for traits with low heritability. At an technical superiority of the final variety. For seed-propa advanced stage of inbreeding (i.e., F. and F7), the best lines gated varieties, it must be feasible to produce seed easily and or mixtures of phenotypically similar lines are typically economically. tested for potential release as new varieties. 25 In addition to phenotypic observations, a plant can also be Mass and recurrent selections can be used to improve characterized by its genotype. The genotype of a plant can populations of either self- or cross-pollinating crops. A be determined by a molecular marker profiling, which can be genetically variable population of heterozygous individuals applied to plants of the same variety or a related variety, can is either identified or created by intercrossing several dif reveal genetic difference of plants and plant parts which are ferent parents. The best plants are selected based on indi- 30 vidual Superiority, outstanding progeny, or excellent com genetically Superior as a result of an event comprising a bining ability. The selected plants are intercrossed to backcross conversion, transgene, or genetic sterility factor, produce a new population in which further cycles of selec and can be used to reveal or validate a pedigree or genetic tion are continued. relationship among test materials. Such molecular marker The single-seed descent procedure in the strict sense 35 profiling can be accomplished by using a variety of tech refers to planting a segregating population, harvesting a niques including, but not limited to, restriction fragment sample of one seed per plant, and using the one-seed sample length polymorphism (RFLP), amplified fragment length to plant the next generation. When the population has been polymorphism (AFLP), sequence-tagged sites (STS), ran advanced from the F to the desired level of inbreeding, the domly amplified polymorphic DNA (RAPD), arbitrarily plants from which lines are derived will each trace to 40 primed polymerase chain reaction (AP-PCR), DNA ampli different F individuals. The number of plants in a popula fication fingerprinting (DAF), sequence characterized tion declines each generation due to failure of some seeds to amplified regions (SCARs), variable number tandem repeat germinate or some plants to produce at least one seed. As a (VNTR), short tandem repeat (STR), single feature poly result, not all of the F plants originally sampled in the morphism (SFP), simple sequence length polymorphism population will be represented by a progeny when genera- 45 (SSLP), restriction site associated DNA, allozymes, isozyme tion advance is completed. markers, single nucleotide polymorphisms (SNPs), or The modified single seed descent procedures involve simple sequence repeat (SSR) markers, also known as harvesting multiple seed (i.e., a single lock or a simple boll) microsatellites (Gupta et al., 1999; Korzun et al., 2001). from each plant in a population and combining them to form Various types of these marker platforms, for example, can be a bulk. Part of the bulk is used to plant the next generation 50 used to identify individual varieties developed from specific and part is put in reserve. This procedure has been used to parent varieties, as well as cells, or other plant parts thereof. save labor at harvest and to maintain adequate seed quan See, for example, Tyagi et al. (2014) “Genetic diversity and tities of the population. The multiple-seed procedure may be population structure in the US Upland cotton (Gossypium used to save labor. It is considerably faster to gin bolls with hirsutum L.). Theoretical and Applied Genetics 127(2): a machine than to remove one seed by hand for the single- 55 283-295; Tatineni et al. (1996) “Genetic diversity in elite seed procedure. The multiple-seed procedure also makes it cotton germplasm determined by morphological character possible to plant the same number of seeds of a population istics and RAPDs.” CropScience 36(1): 186-192; and Cho each generation of inbreeding. Enough seeds are harvested et al. (2014) “Genome-wide SNP marker panel applicable to to make up for those plants that did not germinate or produce Cotton Genetic diversity test.” Proceedings of the Interna seed. 60 tional Cotton Genome Initiative Conference 201):11, each of Descriptions of other breeding methods that are com which are incorporated by reference herein in their entirety. monly used for different traits and crops can be found in one In some examples, one or more markers may be used to of several reference books (e.g., Allard, In: Principles of examine and/or evaluate genetic characteristics of a cotton plant breeding, John Wiley & Sons, NY, University of variety. Particular markers used for these purposes are not California, Davis, Calif., 50-98, 1960; Simmonds, In: Prin- 65 limited to any particular set of markers and diagnostic ciples of crop improvement, Longman, Inc., NY, 369-399, platforms, but are envisioned to include any type of markers 1979; Sneep and Hendriksen, In: Plant breeding perspec and diagnostic platforms that can provide means for distin US 9,615,530 B2 15 16 guishing varieties. One method of genetic characterization variety at inbred stage is essentially homozygous at all may to use only homozygous loci for cotton variety relevant loci, most loci should have only one type of allele 14R938B2XF. present. In contrast, a genetic marker profile of an F. Primers and PCR protocols for assaying these and other progeny should be the Sum of those parents, e.g., if one markers are disclosed in, for example, CottonGen located on 5 parent was homozygous for allele X at a particular locus, the World Wide Web at cottongen.org. In addition to being and the other parent homozygous for allele Y at that locus, used for identification of cotton variety 14R938B2XF, as then the F progeny will be XY (heterozygous) at that locus. well as plant parts and plant cells of cotton variety Subsequent generations of progeny produced by selection 14R938B2XF, a genetic profile may be used to identify a and breeding are expected to be of genotype XX (homozy cotton plant produced through the use of cotton variety 10 gous), YY (homozygous), or XY (heterozygous) for that 14R938B2XF or to verify a pedigree for progeny plants locus position. When the F plant is selfed or sibbed for produced through the use of cotton variety 14R938B2XF. A successive filial generations, the locus should be either X or genetic marker profile may also be useful in breeding and Y for that position. developing backcross conversions. In addition, plants and plant parts Substantially benefiting In an embodiment, the present invention provides a cotton 15 from the use of variety 14R938B2XF in their development, plant characterized by molecular and physiological data such as variety 14R938B2XF comprising a backcross con obtained from a representative sample of said variety depos version, transgene, or genetic sterility factor, may be iden ited with the American Type Culture Collection (ATCC). tified by having a molecular marker profile with a high Thus, plants, seeds, or parts thereof, having all or essentially percent identity to cotton variety 14R938B2XF. Such a all of the physiological and morphological characteristics of percent identity might be 90%, 91%, 92%, 93%, 94%, 95%, cotton variety 14R938B2XF are provided. Further provided 96%, 97%, 98%, 99%, 99.5% or 99.9% identical to cotton is a cotton plant formed by the combination of the disclosed variety 14R938B2XF. cotton plant or plant cell with another cotton plant or cell and A genotypic profile of variety 14R938B2XF also can be comprising the homozygous alleles of the variety. used to identify essentially derived varieties and other In some examples, a plant, a plant part, or a seed of cotton 25 progeny varieties developed from the use of variety variety 14R938B2XF may be characterized by producing a 14R938B2XF, as well as cells and other plant parts thereof. molecular profile. A molecular profile may include, but is Plants of the invention include any plant having at least not limited to, one or more genotypic and/or phenotypic 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, profile(s). A genotypic profile may include, but is not limited 99.5%, or 99.9% of the markers in the genotypic profile, and to, a marker profile. Such as a genetic map, a linkage map. 30 that retain 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, a trait maker profile, a SNP profile, an SSR profile, a 98%, 99%, 99.5%, or 99.9% of the physiological and genome-wide marker profile, a haplotype, and the like. A morphological characteristics of variety 14R938B2XF when molecular profile may also be a nucleic acid sequence grown under the same conditions. Such plants may be profile, and/or a physical map. A phenotypic profile may developed using markers well known in the art. Progeny include, but is not limited to, a protein expression profile, a 35 plants and plant parts produced using variety 14R938B2XF metabolic profile, an mRNA expression profile, and the like. may be identified, for example, by having a molecular One means of performing genetic marker profiling is marker profile of at least 25%, 30%, 35%, 40%, 45%, 50%, using SSR polymorphisms that are well known in the art. A 55%, 60%, 65%, 70%, 75%, 76%, 77%, 78%, 79%, 80%, marker system based on SSRs can be highly informative in 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, linkage analysis relative to other marker systems, in that 40 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or multiple alleles for a given locus may be present. Another 99.5% genetic contribution from cotton variety advantage of this type of marker is that through use of 14R938B2XF, as measured by either percent identity or flanking primers, collecting more informative SSR data can percent similarity. Such progeny may be further character be relatively easily achieved, for example, by using the ized as being within a pedigree distance of variety polymerase chain reaction (PCR), thereby eliminating the 45 14R938B2XF, such as within 1, 2, 3, 4, or 5 or less cross need for labor-intensive Southern hybridization. PCR detec pollinations to a cotton plant other than variety tion may be performed using two oligonucleotide primers 14R938B2XF, or a plant that has variety 14R938B2XF as a flanking the polymorphic segment of repetitive DNA to progenitor. Unique molecular profiles may be identified with amplify the SSR region. other molecular tools, such as SNPs and RFLPs. Following amplification, genotype of test material 50 The two cotton species commercially grown in the United revealed by each marker can be scored by electrophoresis of States are Gossypium hirsutum, commonly known as short the amplification products. Scoring of marker genotype is Staple or upland cotton and Gossypium barbadense, com based on the size of the amplified fragment, which correlates monly known as extra long staple (ELS) or, in the United to the number of base pairs of the fragment. While variation States, as Pima cotton. Upland cotton fiber is used in a wide in the primer used or in the laboratory procedures can affect 55 array of coarser spin count products. Pima cotton is used in the reported fragment size, relative values should remain finer spin count yarns (50-80) which are primarily used in constant regardless of specific primer or laboratory used. more expensive garments. Other properties of Pima cotton When comparing varieties, it may be beneficial to have all are critical because of fiber end use. profiles performed in the same lab. Primers that can be used Cotton is an important and valuable field crop. Thus, a are publically available and may be found in, for example, 60 continuing goal of plant breeders is to develop stable, high CottonGen (Yu et al., CottonGen: a genomics, genetics and yielding cotton varieties that are agronomically sound. The breeding database for cotton research,” Nucleic Acids reasons for this goal are obviously to maximize the amount Research 42 (D1):D1229-D1236, 2013). and quality of the fiber produced on the land used and to A genotypic profile of cotton variety 14R938B2XF can be Supply fiber, oil and food for animals and humans. To used to identify a plant comprising variety 14R938B2XF as 65 accomplish this goal, the cotton breeder must select and a parent, since Such plants will comprise the same homozy develop plants that have the traits that result in superior gous alleles as variety 14R938B2XF. Because the cotton cultivars. US 9,615,530 B2 17 18 II. IMPROVEMENT OF COTTON VARIETIES necessary to introduce a test of the progeny to determine if the desired characteristic has been successfully transferred. In certain further aspects, the invention provides plants Many traits have been identified that are not regularly modified to include at least a first desired trait. Such plants selected for in the development of a new variety but that can may, in one embodiment, be developed by a plant breeding be improved by backcrossing techniques. A genetic locus technique called backcrossing, wherein essentially all of the conferring the traits may or may not be transgenic. Examples desired morphological and physiological characteristics of a of such traits known to those of skill in the art include, but variety are recovered in addition to a genetic locus trans are not limited to, male sterility, herbicide tolerance, resis ferred into the hybrid via the backcrossing technique. The tance for bacterial, fungal, or viral disease, insect or nema term backcrossing as used herein refers to the repeated 10 tode resistance, male sterility, ease of transformation, resis crossing of a hybrid progeny back to a starting variety into tance to abiotic stresses and improved fiber characteristics. which introduction of the desired trait is being carried out. These genes are generally inherited through the nucleus, but The parental plant which contributes the locus or loci for the may be inherited through the cytoplasm. desired trait is termed the nonrecurrent or donor parent. This Direct selection may be applied where a genetic locus acts terminology refers to the fact that the nonrecurrent parent is 15 as a dominant trait. An example of a dominant trait is the used one time in the backcross protocol and therefore does herbicide tolerance trait. For this selection process, the not recur. progeny of the initial cross are sprayed with the herbicide The parental cotton plant to which the locus or loci from prior to the backcrossing. The spraying eliminates any plants the nonrecurrent parent are transferred is known as the which do not have the desired herbicide tolerance charac recurrent parent as it is used for several rounds in the teristic, and only those plants which have the herbicide backcrossing protocol (Poehlman and Sleper, In: Breeding tolerance gene are used in the Subsequent backcross. This Field Crops, Iowa State University Press, Ames, 1995; process is then repeated for all additional backcross genera Sprague and Dudley, In: Corn and Improvement, 3rd ed., tions. 1988: Fehr, In: Principles of variety development. Theory Many useful traits are those which are introduced by and Technique (Vol 1) and Crop Species Soybean (Vol 2), 25 genetic transformation techniques. Methods for the genetic Iowa State Univ., Macmillian Pub. Co., NY, 360-376, transformation of cotton are known to those of skill in the 1987b; Fehr, In: Soybeans. Improvement, Production and art, (see, e.g. Firoozabady et al., Plant Mol. Biol., 10:105 Uses,’ 2d Ed., Monograph 16:249, 1987). In a typical 116, 1987). For example, broadly applicable plant transfor backcross protocol, the original line of interest (recurrent mation methods which have been described include Agro parent) is crossed to a second variety (nonrecurrent parent) 30 bacterium-mediated transformation, microprojectile that carries the genetic locus to be transferred. The resulting bombardment, electroporation, and direct DNA uptake by progeny from this cross are then crossed again to the protoplasts. recurrent parent and the process is repeated until a cotton Agrobacterium-mediated transfer is a widely applicable plant is obtained wherein essentially all of the desired system for introducing gene loci into plant cells, including morphological and physiological characteristics of the recur 35 cotton. An advantage of the technique is that DNA can be rent parent are recovered in the converted plant, in addition introduced into whole plant tissues, thereby bypassing the to the transferred locus from the nonrecurrent parent. need for regeneration of an intact plant from a protoplast. The backcross process may be accelerated by the use of Modern Agrobacterium transformation vectors are capable genetic markers, such as Simple Sequence Length Polymor of replication in E. coli as well as Agrobacterium, allowing phisms (SSLPs) (Williams et al., Nucleic Acids Res., 40 for convenient manipulations (Klee et al., Bio. Tech., 3(7): 18:6531-6535, 1990), Randomly Amplified Polymorphic 637-642, 1985). Moreover, recent technological advances in DNAs (RAPDs), DNA Amplification Fingerprinting (DAF), vectors for Agrobacterium-mediated gene transfer have Sequence Characterized Amplified Regions (SCARs), Arbi improved the arrangement of genes and restriction sites in trary Primed Polymerase Chain Reaction (AP-PCR), Ampli the vectors to facilitate the construction of vectors capable of fied Fragment Length Polymorphisms (AFLPs) (EP 534 45 expressing various polypeptide coding genes. The vectors 858, specifically incorporated herein by reference in its described have convenient multi-linker regions flanked by a entirety), and Single Nucleotide Polymorphisms (SNPs) promoter and a polyadenylation site for direct expression of (Wang et al., Science, 280:1077-1082, 1998) to identify inserted polypeptide coding genes. Additionally, Agrobac plants with the greatest genetic complement from the recur terium containing both armed and disarmed Tigenes can be rent parent. 50 used for transformation. The selection of a suitable recurrent parent is an important In those plant strains where Agrobacterium-mediated step for a Successful backcrossing procedure. The goal of a transformation is efficient, it is the method of choice because backcross protocol is to add or Substitute one or more new of the facile and defined nature of the gene locus transfer. traits in a variety. To accomplish this, a genetic locus of the The use of Agrobacterium-mediated plant integrating vec recurrent parent is modified or substituted with the desired 55 tors to introduce DNA into plant cells is well known in the locus from the nonrecurrent parent, while retaining essen art (Fraley et al., Bio. Tech., 3(7):629-635, 1985; U.S. Pat. tially all of the rest of the desired genetic, and therefore the No. 5,563,055). One efficient means for transformation of desired physiological and morphological constitution of the cotton in particular is transformation and regeneration of original plant. The choice of the particular nonrecurrent cotton hypocotyl explants following inoculation with Agro parent will depend on the purpose of the backcross; one of 60 bacterium tumefaciens from primary callus development, the major purposes is to add some commercially desirable, embryogenesis, embryogenic callus identification, trans agronomically important trait to the plant. The exact back genic cotton shoot production and the development of crossing protocol will depend on the characteristic or trait transgenic plants, as is known in the art. being altered to determine an appropriate testing protocol. To effect transformation by electroporation, for example, Although backcrossing methods are simplified when the 65 one may employ either friable tissues, such as a suspension characteristic being transferred is a dominant allele, a reces culture of cells or embryogenic callus or alternatively one sive allele may also be transferred. In this instance it may be may transform immature embryos or other organized tissue US 9,615,530 B2 19 20 directly. In this technique, one would partially degrade the line'); and (3) a distinct, fertile inbred with normal cyto cell walls of the chosen cells by exposing them to pectin plasm, carrying a fertility restoring gene ("restorer' line). degrading enzymes (pectolyases) or mechanically wound The CMS line is propagated by pollination with the main tissues in a controlled manner. Protoplasts may also be tainer line, with all of the progeny being male sterile, as the employed for electroporation transformation of plants CMS cytoplasm is derived from the female parent. These (Bates, Mol. Biotechnol., 202): 135-145, 1994: Lazzeri, male sterile plants can then be efficiently employed as the Methods Mol. Biol., 49:95-106, 1995). For example, the female parent in hybrid crosses with the restorer line, generation of transgenic cotyledon-derived protoplasts was without the need for physical emasculation of the male described by Dhir and Widholm in Intl. Patent Appl. Publ. reproductive parts of the female parent. No. WO 92/17598, the disclosure of which is specifically 10 The presence of a male-fertility restorer gene results in the incorporated herein by reference. When protoplasts are used, production of fully fertile F hybrid progeny. If no restorer transformation can also be achieved using methods based on gene is present in the male parent, male-sterile hybrids are calcium phosphate precipitation, polyethylene glycol treat obtained. Examples of male-sterility genes and correspond ment, and combinations of these treatments (see, e.g., ing restorers which could be employed with the plants of the Potrykus et al., Mol. Gen. Genet., 199(2):169-177, 1985; 15 invention are well known to those of skill in the art of plant Omirulleh et al., Plant Mol. Biol., 21(3):415-428, 1993: breeding. Examples of such genes include CMS-D2-2, Fromm et al., Nature, 319(6056):791-793, 1986: Uchimiya CMS-hir, CMS-D8, CMS-D4, and CMS-C1. Fertility can be et al., Mol. Gen. Genet., 204(2):204-207, 1986; Marcotte restored to CMS-D2-2 by the D2 restorer in which the and Bayley, Nature, 335(6189):454-457, 1988). restorer factor(s) was introduced from the genome of G. Microprojectile bombardment is another efficient method harknessii Brandegee (D2-2). Microsporogenesis in both for delivering transforming DNA segments to plant cells. In CMS systems aborts during the premeiotic stage. One domi this method, particles are coated with nucleic acids and nant restorer gene from the D8 restorer was identified to delivered into cells by a propelling force. Exemplary par restore fertility of CMS-D8. The D2 restorer for CMS-D2-2 ticles include those comprised of tungsten, platinum, and also restores the fertility of CMS-D8, CMS-hir, and CMS often, gold. For the bombardment, cells in Suspension are 25 C1. concentrated on filters or solid culture medium. Alterna B. Herbicide Tolerance tively, immature embryos or other target cells may be Numerous herbicide tolerance genes are known and may arranged on solid culture medium. The cells to be bom be employed with the plants of the invention. An example is barded are positioned at an appropriate distance below the a gene conferring tolerance to a herbicide that inhibits the macroprojectile stopping plate. 30 growing point or meristem, Such as imidazolinone or Sulfo Microprojectile bombardment techniques are widely nylurea. Exemplary genes in this category code for mutant applicable, and may be used to transform virtually any plant acetolactate synthase (ALS) and acetohydroxy acid synthase species. The application of microprojectile bombardment for (AHAS) enzymes as described, for example, by Lee et al., the transformation of cotton is described, for example, in EMBO.J., 7:1241, 1988: Gleen et al., Plant Molec. Biology, Rajasekaran et al., Mol. Breed., 2:307-319, 1996. An illus 35 18:1185-1187, 1992; Miki et al., Theor. App. Genet., 80:449, trative embodiment of a method for microprojectile bom 1990. bardment is the Biolistics Particle Delivery System, which Tolerance genes for glyphosate (tolerance conferred by can be used to propel particles coated with DNA or cells mutant 5-enolpyruvylshikimate-3-phosphate synthase through a screen, such as a stainless steel or Nytex screen, (EPSP synthase and aroA genes) and hygromycin B phos onto a Surface covered with target cells. The screen disperses 40 photransferase) and to other phosphono compounds such as the particles so that they are not delivered to the recipient glufosinate (phosphinothricin acetyltransferase (PAT) and cells in large aggregates. It is believed that a screen inter Streptomyces hygroscopicus bialaphos resistance (bar) vening between the projectile apparatus and the cells to be genes) may also be used. For example, U.S. Pat. No. bombarded reduces the size of projectiles aggregate and may 4.940,835 to Shah, et al., discloses the nucleotide sequence contribute to a higher frequency of transformation by reduc 45 of a form of EPSP synthase which can confer glyphosate ing the damage inflicted on the recipient cells by projectiles tolerance. A DNA molecule encoding a mutant aroA gene that are too large. can be obtained under ATCC accession number 39256, and It is understood to those of skill in the art that a locus of the nucleotide sequence of the mutant gene is disclosed in transgenic origin need not be directly transformed into a U.S. Pat. No. 4,769,061 to Comai. A hygromycin B phos plant, as techniques for the production of stably transformed 50 photransferase gene from E. coli which confers resistance to cotton plants that pass single loci to progeny by Mendelian glyphosate in tobacco callus and plants is described in inheritance is well known in the art. Such single loci may Penaloza-Vazquez et al. (Plant Cell Reports, 14:482-487. therefore be passed from parent plant to progeny plants by 1995). European patent application No. 0 333 033 to standard plant breeding techniques that are well known in Kumada et al., and U.S. Pat. No. 4,975,374 to Goodman et the art. Non-limiting examples of traits that may be intro 55 al., disclose nucleotide sequences of glutamine synthetase duced directly or by backcrossing are presented below. genes which confer tolerance to herbicides such as phos A. Male Sterility phinothricin. The nucleotide sequence of a phosphinothricin Malesterility genes can increase the efficiency with which acetyltransferase gene is provided in European application hybrids are made, in that they eliminate the need to physi No. 0 242 246 to Leemans et al. DeGreef et al., (Bio/ cally emasculate the plant used as a female in a given cross. 60 Technology, 7:61, 1989), describe the production of trans Where one desires to employ male-sterility systems, it may genic plants that express chimeric bar genes coding for be beneficial to also utilize one or more male-fertility phosphinothricin acetyltransferase activity. Exemplary restorer genes. For example, where cytoplasmic male Ste genes conferring tolerance to herbicidal phenoxy propionic rility (CMS) is used, hybrid crossing requires three inbred acids and cyclohexanediones. Such as haloxyfop and set lines: (1) a cytoplasmically male-sterile line having a CMS 65 hoxydim are the Acct-S1, Accl-S2 and Acct-S3 genes cytoplasm; (2) a fertile inbred with normal cytoplasm, which described by Marshall et al., (Theor. App. Genet., 83:435, is isogenic with the CMS line for nuclear genes (“maintainer 1992). U.S. Patent Application No: 20030135879 describes US 9,615,530 B2 21 22 isolation of a gene for dicamba monooxygenase (DMO) NPR1/SAI1, and/or increasing salicylic acid production from Psueodmonas maltophilia which is involved in the (Ryals et al., Plant Cell, 8:1809-1819, 1996). conversion of a herbicidal form of the herbicidedicamba to Plant defensins may be used to provide resistance to a non-toxic 3,6-dichlorosalicylic acid and thus may be used fungal pathogens (Thomma et al., Planta, 216:193-202, for producing plants tolerant to this herbicide. 5 2002). Genes are also known for conferring tolerance to herbi Nematode resistance has been described, for example, in cides that inhibit photosynthesis, Such as triazine (psbA and U.S. Pat. No. 6.228,992 and bacterial disease resistance in gS+ genes) and benzonitriles such as bromoxynil (nitrilase U.S. Pat. No. 5,516,671. gene). Przibilla et al., (Plant Cell, 3:169, 1991), describe the D. Insect Resistance transformation of Chlamydomonas with plasmids encoding 10 One example of an insect resistance gene includes a mutant psbA genes. Nucleotide sequences for nitrilase genes Bacillus thuringiensis protein, a derivative thereof, or a are disclosed in U.S. Pat. No. 4,810,648 to Stalker, and DNA synthetic polypeptide modeled thereon. See, for example, molecules containing these genes are available under ATCC Geiser et al., (Gene, 48: 109, 1986), who disclose the cloning Accession Nos. 53435, 67441, and 67442. Cloning and and nucleotide sequence of a Bt Ö-endotoxin gene. More expression of DNA coding for a glutathione S-transferase is 15 over, DNA molecules encoding Ö-endotoxin genes can be described by Hayes et al., (Biochem. J. 285:173, 1992). purchased from the American Type Culture Collection, Protoporphyrinogen oxidase (PPO) is the target of the PPO Manassas, Va., for example, under ATCC Accession Nos. inhibitor class of herbicides; a PPO-inhibitor resistant PPO 40098, 67136, 31995 and 31998. Another example is a gene was recently identified in Amaranthus tuberculatus lectin. See, for example, Van Damme et al., (Plant Molec. (Patzoldt et al., PNAS, 103(33): 12329-2334, 2006). The Biol., 24:25, 1994), who disclose the nucleotide sequences herbicide methyl viologen inhibits CO assimilation. Foyer of several Clivia miniata mannose-binding lectin genes. A et al. (Plant Physiol., 109:1047-1057, 1995) describe a plant Vitamin-binding protein may also be used. Such as avidin. overexpressing glutathione reductase (GR) which is resis See PCT application US93/06487, the contents of which are tant to methyl viologen treatment. hereby incorporated by reference. This application teaches Siminszky (Phytochemistry Reviews, 5:445-458, 2006) 25 the use of avidin and avidin homologues as larvicides describes plant cytochrome P450-mediated detoxification of against insect pests. multiple, chemically unrelated classes of herbicides. Yet another insect resistance gene is an enzyme inhibitor, Other examples of herbicide resistance have been for example, a protease or proteinase inhibitor or an amylase described, for instance, in U.S. Pat. Nos. 6,803,501; 6,448, inhibitor. See, for example, Abe et al., J. Biol. Chem., 476; 6,248,876; 6,225,114: 6,107,549; 5,866,775; 5,804, 30 262:16793, 1987 (nucleotide sequence of rice cysteine pro 425; 5,633,435; 5,463,175. teinase inhibitor), Huub et al., Plant Molec. Biol., 21:985, C. Disease Resistance 1993 (nucleotide sequence of cDNA encoding tobacco pro Plant defenses are often activated by specific interaction teinase inhibitor I), and Sumitani et al., Biosci. Biotech. between the product of a disease resistance gene (R) in the Biochem., 57:1243, 1993 (nucleotide sequence of Strepto plant and the product of a corresponding avirulence (AVr) 35 myces nitrosporeus C.-amylase inhibitor). gene in the pathogen. A plant line can be transformed with An insect-specific hormone or pheromone may also be cloned resistance gene to engineer plants that are resistant to used. See, for example, Hammock et al., (Nature, 344:458, specific pathogen strains. See, for example Jones et al., 1990) disclosing baculovirus expression of cloned juvenile Science, 266:789, 1994 (cloning of the tomato Cf-9 gene for hormone esterase, an inactivator of juvenile hormone, Gade resistance to Cladosporium flavum); Martin et al., Science, 40 and Goldsworthy (Eds. Physiological System in Insects, 262: 1432, 1993 (tomato Pto gene for resistance to Elsevier Academic Press, Burlington, Mass., 2007), describ Pseudomonas Syringae pv.); Mindrinos et al., Cell, 78: 1089, ing allostatins and their potential use in pest control; and 1994 (Arabidopsis RPS2 gene for resistance to Pseudomo Palli et al. (Vitam. Horm., 73:59-100, 2005), disclosing use nas Syringae). Logemann et al., (Bio/technology, 10:305. of ecdysteroid and ecdysteroid receptor in agriculture. The 1992), for example, disclose transgenic plants expressing a 45 diuretic hormone receptor (DHR) was identified in Price et barley ribosome-inactivating gene have an increased resis al. (Insect Mol. Biol., 13:469-480, 2004) as a candidate tance to fungal disease. target of insecticides. A viral-invasive protein or a complex toxin derived there Still other examples include an insect-specific antibody or from may also be used for viral disease resistance. For an immunotoxin derived therefrom and a developmental example, the accumulation of viral coat proteins in trans 50 arrestive protein. See Taylor et al., (Seventh Int’l Sympo formed plant cells imparts resistance to viral infection and/or sium on Molecular Plant-Microbe Interactions (Edinburgh, disease development effected by the virus from which the Scotland) Abstract #497, 1994), who described enzymatic coat protein gene is derived, as well as by related viruses. inactivation in transgenic tobacco via production of single See Beachy et al., Ann. Rev. Phytopathol., 28:451, 1990. chain antibody fragments. Coat protein-mediated resistance has been conferred upon 55 E. Resistance to Abiotic Stresses transformed plants against alfalfa mosaic virus, cucumber Abiotic stress includes dehydration or other osmotic mosaic virus, tobacco streak virus, potato virus X, potato stress, salinity, high or low light intensity, high or low virus Y, tobacco etch virus, tobacco rattle virus and tobacco temperatures, Submergence, exposure to heavy metals, and mosaic virus. oxidative stress. Delta-pyrroline-5-carboxylate synthetase A virus-specific antibody may also be used. See, for 60 (P5CS) from mothbean has been used to provide protection example, Tavladoraki et al., (Nature, 366:469, 1993), who against general osmotic stress. Mannitol-1-phosphate dehy show that transgenic plants expressing recombinant anti drogenase (mt1D) from E. coli has been used to provide body genes are protected from virus attack. Additional protection against drought and salinity. Choline oxidase means of inducing whole-plant resistance to a pathogen (codA from Arthrobactor globiformis) can protect against include modulation of the systemic acquired resistance 65 cold and salt. E. coli choline dehydrogenase (betA) provides (SAR) or pathogenesis related (PR) genes, for example protection against salt. Additional protection from cold can genes homologous to the Arabidopsis thaliana NIM1/ be provided by omega-3-fatty acid desaturase (fad7) from US 9,615,530 B2 23 24 Arabidopsis thaliana. Trehalose-6-phosphate synthase and acteristics such as strength, length, fiber fineness, fiber levansucrase (Sacb) from yeast and Bacillus subtilis, maturity ratio, immature fiber content, fiber uniformity, and respectively, can provide protection against drought (Sum micronaire. Cotton plants comprising one or more genes marized from Annex II Genetic Engineering for Abiotic coding for an enzyme selected from the group consisting of Stress Tolerance in Plants, Consultative Group On Interna endoxyloglucan transferase, catalase and peroxidase for the tional Agricultural Research Technical Advisory Commit improvement of cotton fiber characteristics are also tee). Overexpression of Superoxide dismutase can be used to described in U.S. Pat. No. 6,563,022. Cotton modification protect against superoxides, as described in U.S. Pat. No. using ovary-tissue transcriptional factors preferentially 5,538,878 to Thomas et al. directing gene expression in ovary tissue, particularly in F. Modified Fatty Acid, Phytate and Carbohydrate 10 Metabolism very early fruit development, utilized to express genes Genes may be used conferring modified fatty acid encoding isopentenyl transferase in cotton ovule tissue and metabolism. For example, Stearyl-ACP desaturase genes modify the characteristics of boll set in cotton plants and may be used. See Knutzon et al., Proc. Natl. Acad. Sci. USA, alter fiber quality characteristics including fiber dimension 89:2624, 1992. Various fatty acid desaturases have also been 15 and strength is discussed in U.S. Pat. No. 6,329.570. A gene described, such as a Saccharomyces cerevisiae OLE1 gene controlling the fiber formation mechanism in cotton plants is encoding delta-9 fatty acid desaturase, an enzyme which described in U.S. Pat. No. 6,169,174. forms the monounsaturated palmitoleic (16:1) and oleic Genes involved in lignin biosynthesis are described by (18:1) fatty acids from palmitoyl (16:0) or stearoyl (18:0) Dwivedi et al., Mol. Biol., 26:61-71, 1994; Tsai et al., CoA (McDonough et al., J. Biol. Chem., 267(9):5931-5936, Physiol., 107:1459, 1995: U.S. Pat. No. 5,451,514 (claiming 1992); a gene encoding a Stearoyl-acyl carrier protein A9 the use of cinnamyl alcohol dehydrogenase gene in an desaturase from castor (Fox et al. Proc. Natl. Acad. Sci. antisense orientation Such that the endogenous plant cin USA, 90(6):2486-2490, 1993); A6 and A12 desaturases from namyl alcohol dehydrogenase gene is inhibited). the cyanobacteria Synechocystis responsible for the conver H. Additional Traits sion of linoleic acid (18:2) to gamma-linolenic acid (18:3 25 Additional traits can be introduced into the cotton variety gamma) (Reddy et al. Plant Mol. Biol., 22(2):293-300, of the present invention. A non-limiting example of Such a 1993); a gene from Arabidopsis thaliana that encodes an trait is a coding sequence which decreases RNA and/or omega-3 desaturase (Arondel et al. Science, 258(5086): protein levels. The decreased RNA and/or protein levels may 1353-1355, 1992); plant A9 desaturases (PCT Application beachieved through RNAi methods, such as those described Publ. No. WO 91/13972) and soybean and Brassica A15 30 in U.S. Pat. No. 6,506,559 to Fire and Mellow. desaturases (European Patent Application Publ. No. EP Another trait that may find use with the cotton variety of 0616644). the invention is a sequence which allows for site-specific Phytate metabolism may also be modified by introduction recombination. Examples of Such sequences include the of a phytase-encoding gene to enhance breakdown of FRT sequence, used with the FLP recombinase (Zhu and phytate, adding more free phosphate to the transformed 35 Sadowski, J. Biol. Chem., 270:23044-23054, 1995); and the plant. For example, see Van Hartingsveldt et al., (Gene, LOX sequence, used with CRE recombinase (Sauer, Mol. 127:87, 1993), for a disclosure of the nucleotide sequence of Cell. Biol. 7:2087-2096, 1987). The recombinase genes can an Aspergillus niger phytase gene. This, for example, could be encoded at any location within the genome of the cotton be accomplished by cloning and then reintroducing DNA plant, and are active in the hemizygous state. associated with the single allele which is responsible for 40 It may also be desirable to make cotton plants more mutants characterized by low levels of phytic acid. See tolerant to or more easily transformed with Agrobacterium Raboy et al., (Maydica, 35:383, 1990). tumefaciens. Expression of p53 and iap, two baculovirus A number of genes are known that may be used to alter cell-death Suppressor genes, inhibited tissue necrosis and carbohydrate metabolism. For example, plants may be trans DNA cleavage. Additional targets can include plant-encoded formed with a gene coding for an enzyme that alters the 45 proteins that interact with the Agrobacterium Vir genes; branching pattern of starch. See Shiroza et al., J. Bacteol., enzymes involved in plant cell wall formation; and histones, 170:810, 1988 (nucleotide sequence of Streptococcus histone acetyltransferases and histone deacetylases (re mutans fructosyltransferase gene), Steinmetz et al., Mol. viewed in Gelvin, Microbiology & Mol. Biol. Reviews, Gen. Genet., 20:220, 1985 (nucleotide sequence of Bacillus 67:16-37, 2003). subtilis levansucrase gene), Pen et al., BioTechnology, 50 10:292, 1992 (production of transgenic plants that express III. TISSUE CULTURES AND IN VITRO Bacillus licheniformis C.-amylase), Elliot et al., Plant Molec. REGENERATION OF COTTON PLANTS Biol., 21:515, 1993 (nucleotide sequences of tomato inver tase genes), Sergaard et al., J. Biol. Chem., 268:22480, 1993 A further aspect of the invention relates to tissue cultures (site-directed mutagenesis of barley C.-amylase gene), and 55 of the cotton variety designated 14R938B2XF. As used Fisher et al., Plant Physiol., 102:1045, 1993 (maize herein, the term “tissue culture' indicates a composition endosperm starch branching enzyme II). The Z10 gene comprising isolated cells of the same or a different type or encoding a 10 kD Zein storage protein from maize may also a collection of Such cells organized into parts of a plant. be used to alter the quantities of 10 kD Zein in the cells Exemplary types of tissue cultures are protoplasts, calli and relative to other components (Kirihara et al., Mol. Gen. 60 plant cells that are intact in plants or parts of plants, such as Genet., 211:477-484, 1988). embryos, pollen, flowers, leaves, roots, root tips, anthers, G. Improved Cotton Fiber Characteristics and the like. In one embodiment, the tissue culture com Fiber characteristics such as fiber quality of quantity prises embryos, protoplasts, meristematic cells, pollen, represent another example of a trait that may be modified in leaves or anthers. cotton varieties. For example, U.S. Pat. No. 6,472,588 65 An important ability of a tissue culture is the capability to describes transgenic cotton plants transformed with a regenerate fertile plants. This allows, for example, transfor Sucrose phosphate synthase nucleic acid to alter fiber char mation of the tissue culture cells followed by regeneration of US 9,615,530 B2 25 26 transgenic plants. For transformation to be efficient and Desired Agronomic Characteristics: Agronomic charac successful, DNA must be introduced into cells that give rise teristics (which will vary from crop to crop and plant to to plants or germ-line tissue. plant) Such as yield, maturity, pest resistance and lint percent Plants typically are regenerated via two distinct processes; which are desired in a commercially acceptable crop or shoot morphogenesis and somatic embryogenesis. Shoot plant. For example, improved agronomic characteristics for morphogenesis is the process of shoot meristem organiza cotton include yield, maturity, fiber content and fiber quali tion and development. Shoots grow out from a source tissue ties. and are excised and rooted to obtain an intact plant. During Diploid: A cell or organism having two sets of chromo Somatic embryogenesis, an embryo (similar to the Zygotic SOS. embryo), containing both shoot and root axes, is formed 10 Disease Resistance: The ability of plants to restrict the from Somatic plant tissue. An intact plant rather than a activities of a specified pest, such as an insect, fungus, virus, rooted shoot results from the germination of the Somatic embryo. or bacteria. Shoot morphogenesis and somatic embryogenesis are Disease Tolerance: The ability of plants to endure a different processes and the specific route of regeneration is 15 specified pest (such as an insect, fungus, virus or bacteria) or primarily dependent on the explant source and media used an adverse environmental condition and still perform and for tissue culture manipulations. While the systems are produce in spite of this disorder. different, both systems show variety-specific responses Donor Parent: The parent of a variety which contains the where some lines are more responsive to tissue culture gene or trait of interest which is desired to be introduced into manipulations than others. A line that is highly responsive in a second variety. shoot morphogenesis may not generate many Somatic ELS: The abbreviation for “Extra Long Staple.” ELS is embryos. Lines that produce large numbers of embryos the group classification for cotton in the longest staple length during an induction step may not give rise to rapidly category. As used in practice and for commerce, ELS growing proliferative cultures. Therefore, it may be desired denotes varieties belonging to the species G. barbadense to optimize tissue culture conditions for each cotton line. 25 that have Superior fiber qualities, including classification in These optimizations may readily be carried out by one of the longest staple length category. skill in the art of tissue culture through small-scale culture Emasculate: The removal of plant male sex organs or the studies. In addition to line-specific responses, proliferative inactivation of the organs with a cytoplasmic or nuclear cultures can be observed with both shoot morphogenesis and genetic factor conferring male sterility or a chemical agent. somatic embryogenesis. Proliferation is beneficial for both 30 Essentially all of the physiological and morphological systems, as it allows a single, transformed cell to multiply to characteristics: A plant having essentially all of the physi the point that it will contribute to germ-line tissue. ological and morphological characteristics of a designated Embryogenic cultures can also be used Successfully for regeneration, including regeneration of transgenic plants, if plant has all of the characteristics of the plant that are the origin of the embryos is recognized and the biological 35 otherwise present when compared in the same environment, limitations of proliferative embryogenic cultures are under other than an occasional variant trait that might arise during stood. Biological limitations include the difficulty in devel backcrossing or direct introduction of a transgene. oping proliferative embryogenic cultures and reduced fer F Hybrid: The first generation progeny of the cross of tility problems (culture-induced variation) associated with two nonisogenic plants. plants regenerated from long-term proliferative embryo 40 Fallout (Fo): As used herein, the term “fallout” refers to genic cultures. Some of these problems are accentuated in the rating of how much cotton has fallen on the ground at prolonged cultures. The use of more recently cultured cells harvest. may decrease or eliminate Such problems. 2.5% Fiber Span Length: Refers to the longest 2.5% of a bundle of fibers expressed in inches as measured by a digital IV. DEFINITIONS 45 fibergraph. Fiber Characteristics: Refers to fiber qualities such as In the description and tables herein, a number of terms are strength, fiber length, micronaire, fiber elongation, unifor used. In order to provide a clear and consistent understand mity of fiber and amount of fiber. ing of the specification and claims, the following definitions Fiber Elongation: Sometimes referred to as E1, refers to are provided: 50 the elongation of the fiber at the point of breakage in the A: When used in conjunction with the word “comprising strength determination as measured by High Volume Instru or other open language in the claims, the words “a” and “an mentation (HVI). denote “one or more.” Fiber Span Length: The distance spanned by a specific Allele: Any of one or more alternative forms of a gene percentage of fibers in a test specimen, where the initial locus, all of which alleles relate to one trait or characteristic. 55 starting point of the Scanning in the test is considered 100 In a diploid cell or organism, the two alleles of a given gene percent as measured by a digital fibergraph. occupy corresponding loci on a pair of homologous chro Fiber Strength: Also referred to as T1, denotes the force OSOS. required to break a bundle of fibers. Fiber strength is Backcrossing: A process in which a breeder repeatedly expressed in millinewtons (min) per tex on a stelometer. crosses hybrid progeny, for example a first generation hybrid 60 Fruiting Nodes: The number of nodes on the main stem (F), back to one of the parents of the hybrid progeny. from which arise branches that bear fruit or boll in the first Backcrossing can be used to introduce one or more single position. locus conversions from one genetic background into Genotype: The genetic constitution of a cell or organism. another. Gin Turnout: Refers to fraction of lint in a machine Crossing: The mating of two parent plants. 65 harvested Sample of seed cotton (lint, seed, and trash). Cross-pollination: Fertilization by the union of two gam Haploid: A cell or organism having one set of the two sets etes from different plants. of chromosomes in a diploid. US 9,615,530 B2 27 28 Linkage: A phenomenon wherein alleles on the same Transgene: A genetic locus comprising a sequence which chromosome tend to segregate together more often than has been introduced into the genome of a cotton plant by expected by chance if their transmission was independent. transformation. Lint Index: The weight of lint per seed in milligrams. Uniformity Ratio: A measure of the relative fiber span Lint Percent: Refers to the lint (fiber) fraction of seed 5 length uniformity of abundle of fibers. The uniformity ratio cotton (lint and seed). is determined by dividing the 50% fiber span length by the Lint Yield: Refers to the measure of the quantity of fiber 2.5% fiber span length. produced on a given unit of land. Presented herein in pounds Vegetative Nodes: The number of nodes from the coty of lint per acre. ledonary node to the first fruiting branch on the main stem Lint/boll: As used herein, the term “lint/boll is the weight 10 of the plant. of lint per boll. V. DEPOSIT INFORMATION Maturity Rating: A visual rating near harvest on the amount of open bolls on the plant. The rating range is from A deposit of the cotton variety 14R938B2XF, which is 1 to 5, 1 being early and 5 being late. 15 disclosed herein above and referenced in the claims, was Micronaire: A measure of the fineness of the fiber. Within made with the American Type Culture Collection (ATCC), a cotton cultivar, micronaire is also a measure of maturity. 10801 University Blvd., Manassas, Va. 20110-2209. The Micronaire differences are governed by changes in perimeter date of deposit was May 24, 2016 and the accession number or in cell wall thickness, or by changes in both. Within a for those deposited seeds of cotton variety 14R938B2XF is variety, cotton perimeter is fairly consistent and maturity ATCC Accession No. PTA-123144. All restrictions upon the will cause a change in micronaire. Consequently, micronaire deposit have been removed, and the deposit is intended to has a high correlation with maturity within a variety of meet all of the requirements of 37 C.F.R. S.1.801-1.809. The cotton. Maturity is the degree of development of cell wall deposit will be maintained in the depository for a period of thickness. Micronaire may not have a good correlation with 30 years, or 5 years after the last request, or for the effective maturity between varieties of cotton having different fiber 25 life of the patent, whichever is longer, and will be replaced perimeter. Micronaire values range from about 2.0 to 6.0: if necessary during that period. Phenotype: The detectable characteristics of a cell or The references cited herein, to the extent that they provide organism, which characteristics are the manifestation of exemplary procedural or other details Supplementary to gene expression. those set forth herein, are specifically incorporated herein by Plant Height: The average height in meters of a group of 30 reference. plants. What is claimed is: Quantitative Trait Loci (QTL): Quantitative trait loci 1. A plant of cotton variety 14R938B2XF, wherein a (QTL) refer to genetic loci that control to some degree sample of seed of said variety has been deposited under numerically representable traits that are usually continu 35 ATCC Accession No. PTA-123144. ously distributed. 2. A plant part of the plant of claim 1, wherein the plant Recurrent Parent: The repeating parent (variety) in a part comprises at least one cell of said plant. backcross breeding program. The recurrent parent is the 3. The plant part of claim 2, further defined as pollen, a variety into which a gene or trait is desired to be introduced. meristem, a cell, or an ovule. Regeneration: The development of a plant from tissue 40 4. A seed of cotton variety 14R938B2XF, wherein a culture. sample of seed of said variety has been deposited under Seed/bol: Refers to the number of seeds per boll. ATCC Accession No. PTA-123144. Seedcotton/boll: Refers to the weight of seedcotton per 5. A cotton plant that expresses all of the physiological boll. and morphological characteristics of cotton variety Seedweight: Refers to the weight of 100 seeds in grams. 45 14R938B2XF, wherein a sample of seed of said variety has Self-pollination: The transfer of pollen from the anther to been deposited under ATCC Accession No. PTA-123144. the Stigma of the same plant or a plant of the same genotype. 6. A method of producing cotton seed, wherein the Single Locus Converted (Conversion) Plant: Plants which method comprises crossing the plant of claim 1 with itselfor are developed by a plant breeding technique called back a second, distinct cotton plant. crossing wherein essentially all of the desired morphological 50 7. The method of claim 6, wherein the method comprises and physiological characteristics of a variety are recovered crossing the plant of cotton variety 14R938B2XF with a in addition to the characteristics conferred by the single second, distinct cotton plant to produce an F1 hybrid cotton locus transferred into the variety via the backcrossing tech seed. nique. A single locus may comprise one gene, or in the case 8. An Fhybrid cotton seed produced by the method of of transgenic plants, one or more transgenes integrated into 55 claim 7. the host genome at a single site (locus). 9. An Fhybrid cotton plant produced by growing the seed Stringout Rating: also sometimes referred to as “Storm of claim 8. Resistance” refers to a visual rating prior to harvest of the 10. A composition comprising a seed of cotton variety relative looseness of the seed cotton held in the boll structure 14R938B2XF comprised in plant seed growth media, on the plant. The rating values are from 1 to 5 (tight to loose 60 wherein a sample of seed of said variety has been deposited in the boll). under ATCC Accession No. PTA-123144. Substantially Equivalent: A characteristic that, when com 11. The composition of claim 10, wherein the growth pared, does not show a statistically significant difference media is soil or a synthetic cultivation medium. (e.g., p=0.05) from the mean. 12. A plant of cotton variety 14R938B2XF further com Tissue Culture: A composition comprising isolated cells 65 prising a single locus conversion, wherein a sample of seed of the same or a different type or a collection of such cells of cotton variety 14R938B2XF has been deposited under organized into parts of a plant. ATCC Accession No. PTA-123144. US 9,615,530 B2 29 30 13. The plant of claim 12, wherein the single locus (b) growing a progeny plant of a subsequent generation conversion comprises a transgene. from said seed of a progeny plant of a subsequent 14. A seed that produces the plant of claim 12. generation and crossing the progeny plant of a subse 15. The seed of claim 14, wherein the single converted quent generation with itself or a second plant to pro locus confers a trait selected from the group consisting of 5 duce a progeny plant of a further subsequent genera male sterility, herbicide tolerance, insect or pest resistance, tion; and disease resistance, modified fatty acid metabolism, abiotic (c) repeating steps (a) and (b) using said progeny plant of stress resistance, a sequence which allows for site-specific a further subsequent generation from step (b) in place genetic recombination, modified carbohydrate metabolism of the plant grown from said F hybrid cotton seed in and modified cotton fiber characteristics. 10 step (a), wherein steps (a) and (b) are repeated with 16. The seed of claim 15, wherein the single converted Sufficient inbreeding to produce an inbred cotton plant locus confers tolerance to an herbicide selected from the derived from the cotton variety 14R938B2XF. group consisting of glyphosate, sulfonylurea, imidazoli 20. The method of claim 19, further comprising crossing none, dicamba, glufosinate, phenoxy propionic acid, cyclo said inbred cotton plant derived from the cotton variety hexanedione, triazine, benzonitrile, and bromoxynil. 15 14R938B2XF with a plant of a different genotype to produce 17. The seed of claim 15, wherein the trait is insect a seed of a hybrid cotton plant derived from the cotton resistance and said single converted locus comprises a variety 14R938B2XF. transgene encoding a Bacillus thuringiensis (Bt) endotoxin. 21. A method of producing a commodity plant product 18. The seed of claim 15, wherein the single locus comprising collecting the commodity plant product from a comprises a transgene. plant of cotton variety 14R938B2XF, wherein a sample of 19. The method of claim 7, wherein the method further seed of said variety has been deposited under ATCC Acces comprises: Sion No. PTA-123144. (a) crossing a plant grown from said Fhybrid cotton seed 22. The method of claim 21, wherein the commodity plant with itself or a different cotton plant to produce a seed product is lint, seed oil, or seed. of a progeny plant of a subsequent generation;