Us 2018 / 0346920 A1

US 20180346920A1 ( 19) United States (12 ) Patent Application Publication (10 ) Pub. No. : US 2018 /0346920 A1 Nandakumar et al. (43 ) Pub . Date : Dec . 6 , 2018 ( 54 ) MUTATIONS CONFERRING ACETYL -COA (52 ) U . S. CI. CARBOXYLASE ( ACC ) INHIBITING CPC ...... C12N 15 / 8274 ( 2013 .01 ) ; C12N 9 / 93 HERBICIDE TOLERANCE IN SORGHUM ( 2013 .01 ); C12Y 604 / 01002 ( 2013 .01 ) ; A01H ( 71 ) Applicant: Chromatin , Inc ., Chicago , IL (US ) 6 /4666 (2018 .05 ); C12N 15 /8206 (2013 . 01 ) (72 ) Inventors: Rangaraj Nandakumar, Lubbock , TX (US ) ; Song Luo , Chicago , IL (US ) ; (57 ) ABSTRACT Scott A . Staggenborg , Lubbock , TX (US ) The invention provides for sorghum plants and plant parts developed through tissue culture , gene editing or other ( 21) Appl. No. : 15 /993 ,081 methods of mutagenesis in which the plant or plant parts (22 ) Filed : May 30 , 2018 have increased tolerance to one or more acetyl- CoA car boxylase (ACC ) herbicides at levels that would normally Related U .S . Application Data inhibit the growth of wild - type sorghum plants . In this ( 60 ) Provisional application No. 62/ 513 , 074 , filed on May context, the sorghum plantmay be tolerant to any herbicide 31 , 2017 capable of inhibiting acetyl- CoA carboxylase enzyme activ ity . The present invention allows for the screening of ACC Publication Classification herbicide tolerant hybrids with markers or application of (51 ) Int . CI. ACC inhibiting herbicides, and for the removal of unwanted C12N 15 /82 (2006 .01 ) vegetation with application of ACC inhibiting herbicides C12N 9 / 00 ( 2006 .01 ) from seed and grain production fields. AOIH 6 / 46 ( 2006 .01 ) Specification includes a Sequence Listing . Patent Application Publication Dec. 6 , 2018 Sheet 1 of 22 US 2018 / 0346920 A1

BTX430 control calli

......

SO FOP- 13 : W1999C mutant calliLT 3214(BTX430 ) Figure 1 . Patent Application Publication Dec . 6 , 2018 Sheet 2 of 22 US 2018 /0346920 A1

Control

BTX430 Control W1999C mutant calli (BTX430 ) Figure 2 . Patent Application Publication Dec . 6 , 2018 Sheet 3 of 22 US 2018 /0346920 A1

Control plants (BTX430 )

W1999C mutant plants Figure 3 . Patent Application Publication Dec. 6 , 2018 Sheet 4 of 22 US 2018 / 0346920 A1

Figure 4 . DNA and protein sequence data SEQ ID No. 1 . ACC Carboxyl Transferase domain sequence (Wild Type BTX430 ) . GCAAACTCTGGTGCTAGGATTGGCATAGCTGATGAAGTAAAATCTTGCTTC CGTGTTGGGTGGTCTGACGAAGGCAGCCCTGAGCGAGGGTTTCAGTACAT CTATCTGACTGAAGAAGACTATGCCCGTATTAGCTCTTCTGTTATAGCACAT

AAGCTGCAGCTAGATAGCGGTGAAATTAGGTGGATTATTGACTCTGTTGTGTI LAYA1A 1 GGCAAGGAGGATGGGCTTGGTGTTGAGAACATACATGGAAGTGCTGCTAT CGCCAGTGCTTATTCTAGGGCATATGAGGAGACATTTACACTTACATTTGT GACCGGACGGACTGTAGGAATAGGAGCTTATCTTGCTAGACTTGGTATACG GTGCATACAGCGTCTTGACCAGCCGATTATTTTAACAGGGTTTTCTGCCCT GAACAAGCTCCTTGGGCGGGAAGTGTACAGCTCCCACATGCAGCTTGGTGLY GTCCTAAGATCATGGCGACCAATGGTGTTGTCCACCTGACTGTTCCAGATG ACCTTGAAGGTGTTTCCAATATATTGAGGTGGCTCAGCTATGTTCCTGCAA ACATTGGTGGACCTCTTCCTATTACCAAACCTTTGGACCCTCCAGACAGAC CTGTTGCATACATCCCTGAGAACACATGCGATCCACGTGCAGCCATCCGTG GTGTAGATGACAGCCAAGGGAAATGGTTGGGTGGTATGTTTGACAAAGACA GCTTTGTGGAGACATTTGAAGGATGGGCAAAAACAGTGGTTACTGGCAGAG CAAAGCTTGGAGGAATTCCTGTGGGTGTCATAGCTGTGGAGACACAGACCA TGATGCAGCTTGTCCCTGCTGATCCAGGTCAGCTTGATTCCCATGAGCGAT CCGTTCCTCGGGCTGGACAAGTGTGGTTCCCAGATTCTGCAACCAAGACAG CTCAGGCATTATTAGACTTCAACCGTGAAGGATTGCCTCTGTTTATCCTGG CTAACTGGAGAGGTTTCTCTGGTGGACAGAGAGATCTCTTTGAAGGAATTC TTCAGGCTGGGTCAACAATTGTCGAGAACCTTAGGACATATAATCAGCCTG CGTTTGTCTACATTCCTATGGCTGGAGAGCTTCGTGGAGGAGCTTGGGTTG TGGTCGATAGCAAAATAAATCCAGACCGCATTGAGTGTTATGCTGAGAGGA CTGCCAAAGGTAATGTTCTCGAACCTCAAGGGTTAATTGAAATCAAGTTCA GGTCAGAGGAACTCCAAGACTGTATGGGTAGGCTTGACCCCGAGTTGATAA ATCTGAAAGCAAAACTCCAAGATGTAAAGCATGGAAATGGAAGTCTACCAG ACATAGAATCCCTTCAGAAGAGTATAGAAGCACGTACGAAACAGTTGCTGC CITTATATACCCAGATTGCAATACGGTITGCTGAATTGCATGATACTTCCCT AAGAATGGCAGCTAAAGGCGTGATTAAGAAAGTTGTAGACTGGGAAGAATC Patent Application Publication Dec . 6 , 2018 Sheet 5 of 22 US 2018 /0346920 A1

Figure 4 Cont. ACGCTCTTTCTTCTATAAAAGGCTACGGAGAAGGATCTCTGAAGATGTTCT TGCAAAAGAAATAAGACATATAGTCGGTGACAACTTCACTCACCAATCAGC AATGGAGCTCATCAAGGAATGGTACCTGGCTTCTCCAGCCACAGCAGGAAG CACTGGATGGGATGACGATGATGCATTTGTTGCCTGGAAGGACAGTCCTGA AAACTACAATGGATATATCCAAGAGCTAAGGGCTCAAAAAGTGTCTCAGTC GCTCTCTGATCTCACTGACTCCAGTTCAGATCTACAAGCATTCTCGCAGGG TCTTTCTACGCTATTAGATAAGATGGATCCCTCTCAAAGAGCGAAGTTTGTT CAGGAAGTCAAGAAGGTCCTTGGTTGA

Chromatin mutant sequences SEQ ID No. 2 . ACCI ( TGG to TGC ) GCAAACTCTGGTGCTAGGATTGGCATAGCTGATGAAGTAAAATCTTGCTTC CGTGTTGGGTGGTCTGACGAAGGCAGCCCTGAGCGAGGGTTTCAGTACAT CTATCTGACTGAAGAAGACTATGCCCGTATTAGCTCTTCTGTTATAGCACAT AAGCTGCAGCTAGATAGCGGTGAAATTAGGTGGATTATTGACTCTGTTGTGf GGCAAGGAGGATGGGCTTGGTGTTGAGAACATACATGGAAGTGCTGCTAT CGCCAGTGCTTATTCTAGGGCATATGAGGAGACATTTACACTTACATTTGT GACCGGACGGACTGTAGGAATAGGAGCTTATCTTGCTAGACTTGGTATACG GTGCATACAGCGTCTTGACCAGCCAATTATTTTAACAGGGTTTTCTGCCCTYTTET GAACAAGCTCCTTGGGCGGGAAGTGTACAGCTCCCACATGCAGCTTGGTG GTCCTAAGATCATGGCGACCAATGGTGTTGTCCACCTGACTGTTCCAGATG ACCTTGAAGGTGTTTCCAATATATTGAGGTGGCTCAGCTATGTTCCTGCAA ACATTGGTGGACCTCTTCCTATTACCAAACCTTTGGACCCTCCAGACAGAC CTGTTGCATACATCCCTGAGAACACATGCGATCCACGTGCAGCCATCCGTG GTGTAGATGACAGCCAAGGGAAATGGTTGGGTGGTATGTTTGACAAAGACA GCTTTGTGGAGACATTTGAAGGATGGGCAAAAACAGTGGTTACTGGCAGAGYEN ry CAAAGCTTGGAGGAATTCCTGTGGGTGTCATAGCTGTGGAGACACAGACCA TGATGCAGCTTGTCCCTGCTGATCCAGGTCAGCTTGATTCCCATGAGCGATTI CCGTTCCTCGGGCTGGACAAGTGTGOTTCCCAGATTCTGCAACC CTCAGGCATTATTAGACTTCAACCGTGAAGGATTGCCTCTGTTTATCCTGG 19 CTAACTGGAGAGGTTTCTCTGGTGGACAGAGAGATCTCTTTGAAGGAATTC1 1 l Patent Application Publication Dec . 6 , 2018 Sheet 6 of 22 US 2018 /0346920 A1

Figure 4 Cont.

TTCAGGCTGGGTCAACAATTGTCGAGAACCTTAGGACATATAATCAGCCTG1 . TE W CGTTTGTCTACATTCCTATGGCTGGAGAGCTTCGTGGAGGAGCTTGGGTTG TGGTCGATAGCAAAATAAATCCAGACCGCATTGAGTGTTATGCTGAGAGGA CTGCCAAAGGTAATGTTCTCGAACCTCAAGGGTTAATTGAAATCAAGTTCA GGTCAGAGGAACTCCAAGACTGTATGGGTAGGCTTGACCCCGAGTTGATAA ATCTGAAAGCAAAACTCCAAGATGTAAAGCATGGAAATGGAAGTCTACCAG ACATAGAATCCCTTCAGAAGAGTATAGAAGCACGTACGAAACAGTTGCTGC CTTTATATACCCAGATTGCAATACGGTTTGCTGAATTGCATGATACTTCCCT AAGAATGGCAGCTAAAGGCGTGATTAAGAAAGTTGTAGACTGGGAAGAATC ACGCTCTTTCTTCTATAAAAGGCTACGGAGAAGGATCTCTGAAGATGTTCT TGCAAAAGAAATAAGACATATAGTCGGTGACAACTTCACTCACCAATCAGC AATGGAGCTCATCAAGGAATGGTACCTGGCTTCTCCAGCCACAGCAGGAAG CACTGGATGGGATGACGATGATGCATTTGTTGCCTGGAAGGACAGTCCTGA AAACTACAATGGATATATCCAAGAGCTAAGGGCTCAAAAAGTGTCTCAGTC GCTCTCTGATCTCACTGACTCCAGTTCAGATCTACAAGCATTCTCGCAGGG TCTTTCTACGCTATTAGATAAGATGGATCCCTCTCAAAGAGCGAAGTTTGTT CAGGAAGTCAAGAAGGTCCTTGGTTGA

SEQ ID No. 3 . ACC2 ( TGG to TCG ) GCAAACTCTGGTGCTAGGATTGGCATAGCTGATGAAGTAAAATCTTGCTTC CGTGTTGGGTGGTCTGACGAAGGCAGCCCTGAGCGAGGGTTTCAGTACAT CTATCTGACTGAAGAAGACTATGCCCGTATTAGCTCTTCTGTTATAGCACATAIR AT

mm AAGCTGCAGCTAGATAGCGGTGAAATTAGGTGGATTATTGACTCTGTTGTGTA I JA GGCAAGGAGGATGGGCTTGGTGTTGAGAACATACATGGAAGTGCTGCTAT CGCCAGTGCTTATTCTAGGGCATATGAGGAGACATTTACACTTACATTTGT GACCGGACGGACTGTAGGAATAGGAGCTTATCTTGCTAGACTTGGTATACG GTGCATACAGCGTCTTGACCAGCCAATTATTTTAACAGGGTTTTCTGCCCT GAACAAGCTCCTTGGGCGGGAAGTGTACAGCTCCCACATGCAGCTTGGTG GTCCTAAGATCATGGCGACCAATGGTGTTGTCCACCTGACTGTTCCAGATG ACCTTGAAGGTGTTTCCAATATATTGAGGTGGCTCAGCTATGTTCCTGCAALIIGA WIN! Patent Application Publication Dec. 6 , 2018 Sheet 7 of 22 US 2018 / 0346920 A1

Figure 4 Cont. ACATTGGTGGACCTCTTCCTATTACCAAACCTTTGGACCCTCCAGACAGAC CTGTTGCATACATCCCTGAGAACACATGCGATCCACGTGCAGCCATCCGTG GTGTAGATGACAGCCAAGGGAAATGGTTGGGTGGTATGTTTGACAAAGACA GCTTTGTGGAGACATTTGAAGGATGGGCAAAAACAGTGGTTACTGGCAGAG CAAAGCTTGGAGGAATTCCTGTGGGTGTCATAGCTGTGGAGACACAGACCA TGATGCAGCTTGTCCCTGCTGATCCAGGTCAGCTTGATTCCCATGAGCGAT CCGTTCCTCGGGCTGGACAAGTGICOTTCCCAGATTCTGCAACCAAGACAG CTCAGGCATTATTAGACTTCAACCGTGAAGGATTGCCTCTGTTTATCCTGG CTAACTGGAGAGGTTTCTCTGGTGGACAGAGAGATCTCTTTGAAGGAATTC TTCAGGCTGGGTCAACAATTGTCGAGAACCTTAGGACATATAATCAGCCTG CGTTTGTCTACATTCCTATGGCTGGAGAGCTTCGTGGAGGAGCTTGGGTTG TGGTCGATAGCAAAATAAATCCAGACCGCATTGAGTGTTATGCTGAGAGGA CTGCCAAAGGTAATGTTCTCGAACCTCAAGGGTTAATTGAAATCAAGTTCA GGTCAGAGGAACTCCAAGACTGTATGGGTAGGCTTGACCCCGAGTTGATAA ATCTGAAAGCAAAACTCCAAGATGTAAAGCATGGAAATGGAAGTCTACCAG ACATAGAATCCCTTCAGAAGAGTATAGAAGCACGTACGAAACAGTTGCTGC

CTTTATATACCCAGATTGCAATACGGTTTGCTGAATTGCATGATACTTCCCTOLU AAGAATGGCAGCTAAAGGCGTGATTAAGAAAGTTGTAGACTGGGAAGAATC ACGCTCTTTCTTCTATAAAAGGCTACGGAGAAGGATCTCTGAAGATGTTCT TGCAAAAGAAATAAGACATATAGTCGGTGACAACTTCACTCACCAATCAGC AATGGAGCTCATCAAGGAATGGTACCTGGCTTCTCCAGCCACAGCAGGAAG CACTGGATGGGATGACGATGATGCATTTGTTGCCTGGAAGGACAGTCCTGA AAACTACAATGGATATATCCAAGAGCTAAGGGCTCAAAAAGTGTCTCAGTC GCTCTCTGATCTCACTGACTCCAGTTCAGATCTACAAGCATTCTCGCAGGG TCTTTCTACGCTATTAGATAAGATGGATCCCTCTCAAAGAGCGAAGTTTGTT CAGGAAGTCAAGAAGGTCCTTGGTTGA

SEQ ID No. 4 ACC3 (GCA to GTA ) GCAAACTCTGGTGCTAGGATTGGCATAGCTGATGAAGTAAAATCTTGCTTC CGTGTTGGGTGGTCTGACGAAGGCAGCCCTGAGCGAGGGTTTCAGTACAT

CTATCTGACTGAAGAAGACTATGCCCGTATTAGCTCTTCTGTTATAGCACAT. 12 . . Y L L . + Patent Application Publication Dec . 6 , 2018 Sheet 8 of 22 US 2018 /0346920 A1

Figure 4 Cont. AAGCTGCAGCTAGATAGCGGTGAAATTAGGTGGATTATTGACTCTGTTGTG GAGGATGGGCTTGGTGTTGAGAACATACATGGAAGTGCTGCTAT2 t CGCCAGTGCTTATTCTAGGGCATATGAGGAGACATTTACACTTACATTTGT GACCGGACGGACTGTAGGAATAGGAGCTTATCTTGCTAGACTTGGTATACG GTGCATACAGCGTCTTGACCAGCCAATTATTTTAACAGGGTTTTCTGCCCT GAACAAGCTCCTTGGGCGGGAAGTGTACAGCTCCCACATGCAGCTTGGTG GTCCTAAGATCATGGCGACCAATGGTGTTGTCCACCTGACTGTTCCAGATG ACCTTGAAGGTGTTTCCAATATATTGAGGTGGCTCAGCTATGTTCCTGCAA ACATTGGTGGACCTCTTCCTATTACCAAACCTTTGGACCCTCCAGACAGAC CTGTTGCATACATCCCTGAGAACACATGCGATCCACGTGCAGCCATCCGTG GTGTAGATGACAGCCAAGGGAAATGGTTGGGTGGTATGTTTGACAAAGACA GCTTTGTGGAGACATTTGAAGGATGGGCAAAAACAGTGGTTACTGGCAGAG CAAAGCTTGGAGGAATTCCTGTGGGTGTCATAGCTGTGGAGACACAGACCA TGATGCAGCTTGTCCCTGCTGATCCAGGTCAGCTTGATTCCCATGAGCGAT CCGTTCCTCGGGCTGGACAAGTGTCGTTCCCAGATTCTGRAACCAAGACAG CTCAGGCATTATTAGACTTCAACCGTGAAGGATTGCCTCTGTTTATCCTGG CTAACTGGAGAGGTTTCTCTGGTGGACAGAGAGATCTCTTTGAAGGAATTC TTCAGGCTGGGTCAACAATTGTCGAGAACCTTAGGACATATAATCAGCCTG CGTTTGTCTACATTCCTATGGCTGGAGAGCTTCGTGGAGGAGCTTGGGTTG TGGTCGATAGCAAAATAAATCCAGACCGCATTGAGTGTTATGCTGAGAGGA CTGCCAAAGGTAATGTTCTCGAACCTCAAGGGTTAATTGAAATCAAGTTCA GGTCAGAGGAACTCCAAGACTGTATGGGTAGGCTTGACCCCGAGTTGATAA ATCTGAAAGCAAAACTCCAAGATGTAAAGCATGGAAATGGAAGTCTACCAG ACATAGAATCCCTTCAGAAGAGTATAGAAGCACGTACGAAACAGTTGCTGC CTTTATATACCCAGATTGCAATACGGTTTGCTGAATTGCATGATACTTCCCT AAGAATGGCAGCTAAAGGCGTGATTAAGAAAGTTGTAGACTGGGAAGAATC ACGCTCTTTCTTCTATAAAAGGCTACGGAGAAGGATCTCTGAAGATGTTCT TGCAAAAGAAATAAGACATATAGTCGGTGACAACTTCACTCACCAATCAGC AATGGAGCTCATCAAGGAATGGTACCTGGCTTCTCCAGCCACAGCAGGAAG CACTGGATGGGATGACGATGATGCATTTGTTGCCTGGAAGGACAGTCCTGA AAACTACAATGGATATATCCAAGAGCTAAGGGCTCAAAAAGTGTCTCAGTC Patent Application Publication Dec. 6 , 2018 Sheet 9 of 22 US 2018 / 0346920 A1

Figure 4 Cont. GCTCTCTGATCTCACTGACTCCAGTTCAGATCTACAAGCATTCTCGCAGGG TCTTTCTACGCTATTAGATAAGATGGATCCCTCTCAAAGAGCGAAGTTTGTT CAGGAAGTCAAGAAGGTCCTTGGTTGA SEQ ID No . 5 . ACC4 ( TGG to TCG ) TA GCAAACTCTGGTGCTAGGATTGGCATAGCTGATGAAGTAAAATCTTGCTTCILA TH CGTGTTGGGTGGTCTGACGAAGGCAGCCCTGAGCGAGGGTTTCAGTACATII UITIAL CTATCTGACTGAAGAAGACTATGCCCGTATTAGCTCTTCTGTTATAGCACAT

AAGCTGCAGCTAGATAGCGGTGAAATTAGGTGGATTATTGACTCTGTTGTG14. TUU GGCAAGGAGGATGGGCTTGGTGTTGAGAACATACATGGAAGTGCTGCTAT CGCCAGTGCTTATTCTAGGGCATATGAGGAGACATTTACACTTACATTTGT GACCGGACGGACTGTAGGAATAGGAGCTTATCTTGCTAGACTTGGTATACG GTGCATACAGCGTCTTGACCAGCCAATTATTTTAACAGGGTTTTCTGCCCT GAACAAGCTCCTTGGGCGGGAAGTGTACAGCTCCCACATGCAGCTTGGTG GTCCTAAGATCATGGCGACCAATGGTGTTGTCCACCTGACTGTTCCAGATG ACCTTGAAGGTGTTTCCAATATATTGAGGTGGCTCAGCTATGTTCCTGCAAWIU ACATTGGTGGACCTCTTCCTATTACCAAACCTTTGGACCCTCCAGACAGAC CTGTTGCATACATCCCTGAGAACACATGCGATCCACGTGCAGCCATCCGTG GTGTAGATGACAGCCAAGGGAAATGGTTGGGTGGTATGTTTGACAAAGACA GCTTTGTGGAGACATTTGAAGGATGGGCAAAAACAGTGGTTACTGGCAGAG CAAAGCTTGGAGGAATTCCTGTGGGTGTCATAGCTGTGGAGACACAGACCA TGATGCAGCTTGTCCCTGCTGATCCAGGTCAGCTTGATTCCCATGAGCGAT CCGTTCCTCGGGCTGGACAAGTGTCGTTCCCAGATTCTGTAACCAAGACAG CTCAGGCATTATTAGACTTCAACCGTGAAGGATTGCCTCTGTTTATCCTGG CTAACTCGAGAGGTTTCTCTGGTGGACAGAGAGATCTCTTTGAAGGAATTC TTCAGGCTGGGTCAACAATTGTCGAGAACCTTAGGACATATAATCAGCCTG CGTTTGTCTACATTCCTATGGCTGGAGAGCTTCGTGGAGGAGCTTGGGTTG TGGTCGATAGCAAAATAAATCCAGACCGCATTGAGTGTTATGCTGAGAGGA CTGCCAAAGGTAATGTTCTCGAACCTCAAGGGTTAATTGAAATCAAGTTCA GGTCAGAGGAACTCCAAGACTGTATGGGTAGGCTTGACCCCGAGTTGATAA ATCTGAAAGCAAAACTCCAAGATGTAAAGCATGGAAATGGAAGTCTACCAG Patent Application Publication Dec . 6 , 2018 Sheet 10 of 22 US 2018 /0346920 A1

Figure 4 Cont. ACATAGAATCCCTTCAGAAGAGTATAGAAGCACGTACGAAACAGTTGCTGC CTTTATATACCCAGATTGCAATACGGTTTGCTGAATTGCATGATACTTCCCT AAGAATGGCAGCTAAAGGCGTGATTAAGAAAGTTGTAGACTGGGAAGAATC ACGCTCTTTCTTCTATAAAAGGCTACGGAGAAGGATCTCTGAAGATGTTCT TGCAAAAGAAATAAGACATATAGTCGGTGACAACTTCACTCACCAATCAGC AATGGAGCTCATCAAGGAATGGTACCTGGCTTCTCCAGCCACAGCAGGAAG CACTGGATGGGATGACGATGATGCATTTGTTGCCTGGAAGGACAGTCCTGA AAACTACAATGGATATATCCAAGAGCTAAGGGCTCAAAAAGTGTCTCAGTC GCTCTCTGATCTCACTGACTCCAGTTCAGATCTACAAGCATTCTCGCAGGG TCTTTCTACGCTATTAGATAAGATGGATCCCTCTCAAAGAGCGAAGTTTGTT CAGGAAGTCAAGAAGGTCCTTGGTTGA

SEQ ID . No. 6 Sorghum Wild type CT domain sequence ( Highlighted entries replaced in mutations) ANSGARIGIADEVKSCFRVGWSDEGSPERGFOYIYLTEEDYARISSSVIAHKLQL DSGEIRWIIDSVVGKEDGLGVENIHGSAAIASAYSRAYEETFTLTFVTGRTVGIG AYLARLGIRCIQRLDQPIILTGFSALNKLLGREVYSSHMQLGGPKIMATNGVVH LTVPDDLEGVSNILRWLSYVPANIGGPLPITKPLDPPDRPVAYIPENTCDPRAAI RGVDDSOGKWLGGMFDKDSFVETFEGWAKTVVTGRAKLGGIPVGVIAVETOT MMQLVPADPGQLDSHERSVPRAGOVWFPDSATKTAQALLDFNREGLPLFILAN WRGFSGGQRDLFEGILQAGSTIVENLRTYNQPAFVYIPMAGELRGGAWVVVDS KINPDRIECYAERTAKGNVLEPOGLIEIKFRSEELODCMGRLDPELINLKAKLO DVKHGNGSLPDIESLOKSIEARTKOLLPLYTQIAIRFAELHDTSLRMAAKGVIK KVVDWEESRSFFYKRLRRRISEDVLAKEIRHIVGDNFTHOSAMELIKEWYLASP ATAGSTGWDDDDAFVAWKDSPENYNGYIQELRAQKVSQSLSDLTDSSSDLQAF SQGLSTLLDKMDPSQRAKFVQEVKKVLG *

SEQ ID No. 7 . ACCI (W1999C ) ANSGARIGIADEVKSCFRVGWSDEGSPERGFQYIYLTEEDYARISSSVIAHKLQL DSGEIRWIIDSVVGKEDGLGVENIHGSAAIASAYSRAYEETFTLTFVTGRTVGIG AYLARLGIRCIQRLDQPIILTGFSALNKLLGREVYSSHMQLGGPKIMATNGVVH Patent Application Publication Dec . 6 , 2018 Sheet 11 of 22 US 2018 /0346920 A1

Figure 4 Cont. LTVPDDLEGVSNILRWLSYVPANIGGPLPITKPLDPPDRPVAYIPENTCDPRAAI RGVDDSQGKWLGGMFDKDSFVETFEGWAKTVVTGRAKLGGIPVGVIAVETQT MMQLVPADPGQLDSHERSVPRAGQVCFPDSATKTAQALLDFNREGLPLFILAN WRGFSGGORDLFEGILQAGSTIVENLRTYNQPAFVYIPMAGELRGGAWVVVDS KINPDRIECYAERTAKGNVLEPQGLIEIKFRSEELQDCMGRLDPELINLKAKLQ DVKHGNGSLPDIESLOKSIEARTKQLLPLYTQIAIRFAELHDTSLRMAAKGVIK KVVDWEESRSFFYKRLRRRISEDVLAKEIRHIVGDNFTHOSAMELIKEWYLASP ATAGSTGWDDDDAFVAWKDSPENYNGYIQELRAQKVSQSLSDLTDSSSDLQAF SQGLSTLLDKMDPSQRAKFVQEVKKVLG *

SEQ ID No. 8 . ACC2 (W1999S ) ANSGARIGIADEVKSCFRVGWSDEGSPERGFQYIYLTEEDYARISSSVIAHKLQL DSGEIRWIIDSVVGKEDGLGVENIHGSAAIASAYSRAYEETFTLTFVTGRTVGIG AYLARLGIRCIQRLDOPIILTGFSALNKLLGREVYSSHMQLGGPKIMATNGVVH LTVPDDLEGVSNILRWLSYVPANIGGPLPITKPLDPPDRPVAYIPENTCDPRAAI RGVDDSQGKWLGGMFDKDSFVETFEGWAKTVVTGRAKLGGIPVGVIAVETOT MMQLVPADPGOLDSHERSVPRAGOVSFPDSATKTAQALLDFNRÉGLPLFILAN WRGFSGGQRDLFEGILQAGSTIVENLRTYNQPAFVYIPMAGELRGGAWVVVDS KINPDRIECYAERTAKGNVLEPQGLIEIKFRSEELQDCMGRLDPELINLKAKLQ DVKHGNGSLPDIESLQKSIEARTKQLLPLYTQIAIRFAELHDTSLRMAAKGVIK KVVDWEESRSFFYKRLRRRISEDVLAKEIRHIVGDNFTHOSAMELIKEWYLASP ATAGSTGWDDDDAFVAWKDSPENYNGYIQELRAQKVSQSLSDLTDSSSDLQAF SQGLSTLLDKMDPSQRAKFVQEVKKVLG * SEQ ID No . 9. ACC3 ( A2004V ) ANSGARIGIADEVKSCFRVGWSDEGSPERGFOYIYLTEEDYARISSSVIAHKLQL DSGEIRWIIDSVVGKEDGLGVENIHGSAAIASAYSRAYEETFTLTFVTGRTVGIG AYLARLGIRCIQRLDQPIILTGFSALNKLLGREVYSSHMOLGGPKIMATNGVVH LTVPDDLEGVSNILRWLSYVPANIGGPLPITKPLDPPDRPVAYIPENTCDPRAAI RGVDDSOGKWLGGMFDKDSFVETFEGWAKTVVTGRAKLGGIPVGVIAVETOT MMQLVPADPGQLDSHERSVPRAGQVWFPDSVIKTAQALLDFNREGLPLFILAN Patent Application Publication Dec . 6 , 2018 Sheet 12 of 22 US 2018 /0346920 A1

Figure 4 Cont. WRGFSGGQRDLFEGILQAGSTIVENLRTYNOPAFVYIPMAGELRGGAWVVVDS KINPDRIECYAERTAKGNVLEPQGLIEIKFRSEELQDCMGRLDPELINLKAKLQ DVKHGNGSLPDIESLQKSIEARTKQLLPLYTQIAIRFAELHDTSLRMAAKGVIK KVVDWEESRSFFYKRLRRRISEDVLAKEIRHIVGDNFTHQSAMELIKEWYLASP ATAGSTGWDDDDAFVAWKDSPENYNGYIQELRAQKVSQSLSDLTDSSSDLQAF SQGLSTLLDKMDPSQRAKFVQEVKKVLG *

SEQ ID No. 10 . ACC4 (W20275 ) ANSGARIGIADEVKSCFRVGWSDEGSPERGFQYIYLTEEDYARISSSVIAHKLQL GEIRWIIDSVVGKEDGLGVENIHGSAAIASAYSRAYEETFTLTFVTGRTVGIG AYLARLGIRCIQRLDQPIILTGFSALNKLLGREVYSSHMQLGGPKIMATNGVVH LTVPDDLEGVSNILRWLSYVPANIGGPLPITKPLDPPDRPVAYIPENTCDPRAAI RGVDDSQGKWLGGMFDKDSFVETFEGWAKTVVTGRAKLGGIPVGVIAVETQT MMQLVPADPGQLDSHERSVPRAGQVWFPDSATKTAQALLDFNREGLPLFILAN SRGFSGGORDLFEGILQAGSTIVENLRTYNQPAFVYIPMAGELRGGAWVVVDS KINPDRIECYAERTAKGNVLEPQGLIEIKFRSEELODCMGRLDPELINLKAKLO DVKHGNGSLPDIESLQKSIEARTKQLLPLYTQIAIRFAELHDTSLRMAAKGVIK KVVDWEESRSFFYKRLRRRISEDVLAKEIRHIVGDNFTHQSAMELIKEWYLASP ATAGSTGWDDDDAFVAWKDSPENYNGYIQELRAQKVSQSLSDLTDSSSDLQAF SOGLSTLLDKMDPSORAKFVOEVKKVLG * Patent Application Publication Dec . 6 , 2018 Sheet 13 of 22 US 2018 /0346920 A1

2030SEQIDNO:62030SEQ IDNO:72030 SEQID NO:8 :2030SEOIDNO9 2030SEQIDNO:10

SVPRAGOVWEPDSAIKTAQALLDENREGLPLFILANIANS SVPRAGOVOFPDSTKTAQALLDENREGLPLFILANDANS SVPRAGOVIFPDSVIKTAQALLDENREGLPLEILANLANS SVPRAGOVIFPDSTKTAQALLDENREGLPLEILANSANS Figure5 1667 SVPRAGOVSEPOSEIKTAQALLDENREGLPLEILANLANS1991 1991 1991 Wildtype W1999C W19995 A2004V W20275 Patent Application Publication Dec . 6 , 2018 Sheet 14 of 22 US 2018 /0346920 A1

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F2- P12 (W1999C mutant) Wild type BTX430 Figure 14B US 2018 /0346920 A1 Dec . 6 , 2018

MUTATIONS CONFERRING ACETYL - COA [0008 ] The development of herbicide resistance in plants CARBOXYLASE ( ACC ) INHIBITING offers significant production and economic advantages ; as HERBICIDE TOLERANCE IN SORGHUM such the use of herbicides for controlling weeds or plants in crops has become almost a universal practice . However , [0001 ] This application claim priority to U . S . Provisional application of such herbicides can also result in death or Patent Application No. 62 /513 ,074 , filed May 31 , 2017 , reduced growth of the desired crop plant, making the time which is incorporated by reference herein in its entirety . and method of herbicide application critical or in some cases unfeasible . INCORPORATION BY REFERENCE OF [0009 ] Of particular interest to farmers is the use of SEQUENCE LISTING herbicides with greater potency , broad weed spectrum effec [ 0002 ] This application , as a separate part of disclosure , a tiveness and rapid soil degradation . Plants , plant tissues and Sequence Listing in computer- readable form ( filename: seeds with resistance to these compounds would provide an 52092 _ Seqlisting . txt; 44 , 821 bytes -ASCII text file ; created attractive solution by allowing the herbicides to be used to May 24 , 2018 ) which is incorporated by reference herein in control weed growth , without risk of damage to the crop . its entirety . ACC herbicides are those with the mode of action that affect the Acetyle - CoA carboxylase enzyme in the plant. This class of herbicide is only effective in controlling member of the FIELD OF INVENTION Poaceae or Gramineae family of plants. Such herbicides are [0003 ] The invention provides for sorghum plants and included in the aryloxyphenoxypropionate ( FOP ), cyclo plant parts developed through tissue culture , gene editing or hexanedione (DIM ) and phenylpyrazolin (DENs ) chemical other methods of mutagenesis in which the plant or plant families. For example , sorghum is susceptible to many ACC parts have increased resistance to one or more acetyl- CoA inhibiting herbicides that target monocot species , making carboxylase (ACC ) herbicides at levels that would normally the use of these herbicides to control grassy weeds almost inhibit the growth of wild - type sorghum plants . In this impossible with conventional sorghum hybrids and open context, the sorghum plant may be tolerant to any herbicide pollinated varieties . capable of inhibiting acetyl- CoA carboxylase enzyme activ [0010 ] Certain weed grass species have been found that ity . For example , the sorghum plant may be tolerant to display altered sensitivity to FOP and DIM herbicides . One herbicides of the aryloxyphenoxypropionate ( FOP ) , cyclo grass species , black grass ( A . myosuroides [Huds . ] ) , is a hexanedione (DIM ) and phenylpyrazolin (DENs ) herbicide major grass weed in Europe . Several mutations have been family . This invention allows for creation of ACC herbicide found in the genome of some black grass plants that confer resistant lines and hybrid seed through tissue culture or resistance to some, but not all, FOP and DIM herbicides transgenic methods including gene editing methods, with up (Délve , et al. , 2005 , Plant Phys . 137 : 794 - 806 ; Délve , et al. , to 25 or 50 % efficiencies of regenerating desired plants . The 2002 , Theor . Appl. Genet . 104 : 1114 - 1120 ) . Similar findings present invention allows for the screening of ACC herbicide were found in mutant grass weeds such as annual ryegrass tolerant hybrids with markers or application of ACC inhib ( L . rigidum [Gaud . ] ; Délye , et al. , 2002 , Pest Manag . Sci . iting herbicides, and for the removal of unwanted vegetation 58 :474 -478 ), green foxtail ( S . viridis ( L . Beauv. ) ; Zhang and with application of ACC inhibiting herbicides from seed and Devine , 2000 , Weed Sci. Soc. Am . 40 :33 ; Délye , et al ., 2002 , grain production fields. Planta 214 : 421 -427 ) and wild oat ( A . fatua [ L . ) ; Christoffers et al. , 2002 , Genome 45 : 1049 - 1056 ) . One herbicide resistant BACKGROUND maize hybrid (DK592 from Dekalb ) has a similar mutation in the ACC enzyme as that found in grass weeds (Zagnitko [0004 ] Sorghum is the second most important cereal- feed et al. , 2001, Proc . Natl . Acad . Sci. 98 : 6617 - 22 ) . grain grown in the United States . Production is economi [0011 ] Creating mutations as means of altering a plants cally critical to farms operating in marginal rainfall areas phenotype and composition is a common modern plant because of sorghum ' s ability to tolerate drought and heat. improvement practice . This can be accomplished through Both the livestock and bio - energy industries utilize sorghum chemical or DNA damaging mutagenesis , gene editing , or as an energy substrate thereby making it a versatile crop . through tissue culture selection . Chemical mutagenesis is [ 0005 ] Sorghum is more tolerant to drought and excess soil the process where plant tissue, normally seeds, are exposed moisture content than most cereals . It is capable of growing to a mutagen inducing compounds like Ethy methanesul properly under varied soil and weather conditions . Likewise , fonate ( EMS ) , sodimum Azide ( AZ ) , or methylnitrosoureas it responds favorably to irrigation , requiring a minimum of (MNU ) , or irradiated with X - rays, fast neutrons, or other 250 mm during its life cycle , with an optimum irrigation types of DNA damaging particles . Mutagenized seed is then ranging from 400 - 550 mm . planted and the desired mutation is selected through a [0006 ] Furthermore , sorghum has the ability of remaining variety of methods, such as exposing plants from the muta dormant during periods of drought and resumes growth genized seed to an herbicide for which a resistant or tolerant under favorable periods , although these stress situations may plant is desired . affect performance . [0012 ] Creating mutations via tissue culture occurs by [ 0007 ] Worldwide , sorghum is the fifth leading cereal exposing undifferentiated cells , in callus , to the stress of grain . As it is tolerant to both drought and heat , it is easily choice in gradually increasing intensities to allow mutations the most widely grown food grain in the semiarid regions of to occur in the callus cells. The cells that mutate and survive sub - Sahelian Africa and in the dry central peninsular region the exposure and propagate and are exposed to gradually of India . As such , sorghum is used in human consumption in higher levels of intensity of the stress until a desired level is most of the driest regions of the world thereby making it a achieved . At this point, the callus must be transformed into critically important food crop in these locations . a plant and propagated from seed , in the case of crop plants . US 2018 /0346920 A1 Dec. 6 , 2018

Targeted genome editing is useful for creating plant traits thereby allowing for greater crop yield when these herbi and phenotypes, as well as for plant breeding. Multiple gene cides are used to control grassy weeds . editing technologies were developed in the past years , [0017 ]. The present invention provides for compositions including zinc finger nucleases ( ZFNs) , transcription acti and methods for producing sorghum plants , seeds, and plant vator- like effector nucleases ( TALENs) , and clustered regu parts that have modified acetyl- CoA carboxylase ( ACC ) larly interspaced short palindromic repeat ( CRISPR ) genes and proteins that make these plants , seeds, and plant CRISPR - associated protein (Cas ) systems, such as Cas9 , parts resistant to inhibition by herbicides that normally Cas3 , Cas8a , Cas5 , Cas8b , Cas8c , Cas10d , Csel, Cse2 , inhibit the activity of the ACC protein . Csyn , Csy2 , Csy3 , GSU0054 , Cas 10 , Csm2, Cmr5 , Cas 11 , Csx10 , Csfl , Csn2 , Cas4 , Cpf1 , C2c1 , c2c3 , Cas13a , SUMMARY OF INVENTION Cas13b , and Cas 1. These editing platforms allow for reverse [0018 ] The invention provides for ACC inhibitor herbicide genetics , genome engineering and targeted transgene inte tolerant sorghum plants or plant parts thereof comprising gration by inducing DNA double strands breaks in the one or more mutations of the Acetyl - CoA Carboxylase specific genomic loci of a cell and then harnessing the cell' s (ACC ) gene , wherein the sorghum plant or plant part has natural repair pathways . increased resistance to one or more ACC inhibiting herbi [0013 ] One of the distinct advantages of creating muta cide as compared with a wild - type sorghum cultivar or plant. tions with either mutagenesis , gene editing , or tissue culture The nucleotide sequence of the wild -type (un modified ) is that the trait of interest, herbicide tolerance in the case ACC sorghum gene is set out as SEQ ID NO : 1 . here , can be developed in elite breeding germplasm . This [0019 ] The ACC inhibitor herbicide tolerant sorghum approach results in more stable breeding germplasm than plants or plant parts of the invention include plant or plant selecting for similar mutations in wild relatives as was the part corresponding to the deposit under ATCC Accession case in Tunistra and Al- Khatib ( 2017 ) . No. PTA - 125106 , PTA - 125107 or PTA - 125108 , deposited [ 0014 ] Three significant inventions exist in the area of on May 9 , 2018 with the American Type Tissue Culture sorghum herbicide tolerance . Trucillo et al. Collection (ATCC ) . The invention also provides for plant (WO2013149674A1 , WO2013149674A8) used chemical progeny of these ACC inhibitor herbicide tolerant sorghum mutagenesis to create mutations to create sorghum germ plants . plasm that inhibits AHAS enzyme activity , inferring toler [ 0020 ] The invention also provides for seed corresponding ance to imidazolinone herbicides . Tuinstra and Al- Khatib to the deposit under ATCC Accession No. PTA - 125106 , ( U . S . Pat . No . 9 ,617 , 530 ) created elite sorghum germplasm PTA - 125107 or PTA - 125108 , deposited on May 9 , 2018 that contained altered acetyle - CoA carboxylase ( ACC ) with the American Type Tissue Culture Collection (ATCC ) . genes that inferred resistance to acetyl- CoA carboxylase The invention also provides for plant progeny of resulting herbicides which are herbicides from the aryloxyphenoxy from the seed . propionate (FOP ) and cyclohexanedione (DIM ) chemical (0021 ] In an embodiment of the invention , the ACC families. Tuinstra and Al- Khatib (WO2008073800AS ) also inhibitor herbicide tolerant sorghum plants or plant parts created elite sorghum germplasm that contained altered thereof comprising one or more mutations of the sorghum acetolactate synthase (ALS ) genes and proteins that are ACC gene , wherein the nucleotide sequence encoding the resistant to inhibition by herbicides that normally inhibit the CT domain of the ACC protein comprises one of the activity of the ALS protein . Both Tuinstra inventions were following: the nucleotide sequence of SER ID NO : 2 ; the developed by screening wild sorghum relatives that con nucleotide sequence of SEQ ID NO : 3 ; the nucleotide tained the altered genes , respectively . sequence of SEO ID NO : 4 ; the nucleotide sequence of SEO [0015 ] Acetyl- CoA carboxylase ( ACC ) is a biotinylated ID NO : 5 ; the nucleotide sequence of SEQ ID NO : 2 and one enzyme that catalyzes the carboxylation of acetyl- CoA to of the following : the nucleotide sequence of SEQ ID NO : 3 , produce malonyl - CoA . This carboxylation is a two - step , SEO ID NO : 4 or SEQ ID NO : 5 ; the nucleotide sequence reversible reaction consisting of the ATP - dependent car of SEQ ID NO : 3 and one of the following: the nucleotide boxylation of the biotin group on the carboxyl carrier sequence of SEQ ID NO : 4 or SEQ ID NO : 5 ; the nucleotide domain by biotin - carboxylase activity followed by the trans sequence of SEQ ID NO : 4 and SEQ ID NO : 5 ; the nucleo fer of the carboxyl group from biotin to acetyl- CoA by tide sequence of SEQ ID NO : 2 , SEQ ID NO : 3 and SEQ ID carboxyltransferase activity (Nikolau et al. , 2003 , Arch . NO : 4 ; the nucleotide sequence of SEQ ID NO : 2 , SEQ ID Biochem . Biophys . 414 :211 - 22 ). Acetyl- CoA carboxylase is NO : 3 and SEQ ID NO : 5 ; the nucleotide sequence of SEQ not only a key enzyme in plants for biosynthesis of fatty ID NO : 2 , SEQ ID NO : 4 and SEQ ID NO : 5 ; or The acids , a process that occurs in chloroplasts and mitochon nucleotide sequence of SEQ ID NO : 3 , SEQ ID NO : 4 and dria , but ACC also plays a role in the formation of long SEQ ID NO : 5 and wherein the sorghum plant or plant part chain fatty acids and flavonoids, and in malonylation that has increased resistance to one or more ACC inhibiting occurs in the cytoplasm . There are two isoforms of ACC herbicide as compared with a wild - type sorghum cultivar or with the chloroplastic ACC accounting for more than 80 % of plant. The ACC inhibitor herbicide tolerant sorghum plant the total ACC activity (Herbert et al. , 1996 , Biochem . J . parts may be an organ , tissue, cell or seed . 318 : 997 - 1006 ) . Aryloxyphenoxypropionate (FOP ) and [0022 ] In another embodiment of the invention , the ACC cyclohexanedione (DIM ) are two classes of chemicals that inhibitor herbicide tolerant sorghum plants or plant parts are known to selectively inhibit chloroplastic ACC in thereof comprising one or more mutations of the sorghum grasses (Rendina et al. , 1990 , J . Agric . Food Chem . 38 : 1282 ACC gene , wherein ACC gene encodes a sorghum acetyl 1287 ) . CoA protein having a CT domain comprising one or more of [ 0016 ] Due to the importance of sorghum as a crop plant the following mutations: a Tryptophan to Cysteine amino on the world stage , what are needed are sorghum hybrids acid substitution at an amino acid position 1999 (W1999C ; that are resistant to the inhibitory effects of ACC herbicides, SEQ ID NO : 7 ) aligning with the amino acid sequence of US 2018 /0346920 A1 Dec. 6 , 2018

SEQ ID NO : 6 , or a Tryptophan to Serine amino acid SEQ ID NO : 9 ) aligning with the amino acid sequence of substitution at an amino acid position 1999 (W1999S ; SEQ SEQ ID NO : 6 and a Tryptophan to Serine amino acid ID NO : 8 ) aligning with the amino acid sequence of SEQ ID substitution at an amino acid position 2027 (W2027S ; SEQ NO : 6 , or an alanine to valine amino acid substitution at an ID NO : 10 ) aligning with the amino acid sequence of SEQ amino acid position 2004 ( A2004V ; SEQ ID NO : 9 ) aligning ID NO : 6 , or a Tryptophan to Serine amino acid substitution with the amino acid sequence of SEQ ID NO : 6 , or a at an amino acid position 1999 (W1999S ; SEQ ID NO : 8 ) Tryptophan to Serine amino acid substitution at an amino aligning with the amino acid sequence of SEQ ID NO : 6 and an alanine to valine amino acid substitution at an amino acid acid position 2027 (W2027S ; SEQ ID NO : 10 ) aligning with position 2004 ( A2004V ; SEQ ID NO : 9 ) aligning with the the amino acid sequence of SEQ ID NO : 6 , or a Tryptophan amino acid sequence of SEQ ID NO : 6 and a Tryptophan to to Cysteine amino acid substitution at an amino acid position Serine amino acid substitution at an amino acid position 1999 (W1999C ; SEQ ID NO : 7 ) aligning with the amino 2027 (W2027S ; SEQ ID NO : 10 ) aligning with the amino acid sequence of SEQ ID NO : 6 and a Tryptophan to Serine acid sequence of SEQ ID NO : 6 , wherein the sorghum plant amino acid substitution at an amino acid position 1999 or plant part has increased resistance to one or more ACC (W1999S ; SEQ ID NO : 8 ) aligning with the amino acid inhibiting herbicide as compared with a wild -type sorghum sequence of SEQ ID NO : 6 , or a Tryptophan to Cysteine cultivar or plant. SEQ ID NO : 6 is the amino acid sequence amino acid substitution at an amino acid position 1999 (W1999C ; SEQ ID NO : 7 ) aligning with the amino acid of the wild -type ACC protein CT domain . sequence of SEQ ID NO : 6 and an alanine to valine amino [0023 ] Any of the ACC inhibitor herbicide tolerant sor acid substitution at an amino acid position 2004 (A2004V ; ghum plant or plant parts of the invention are tolerant or SEQ ID NO : 9 ) aligning with the amino acid sequence of resistant to an aryloxyphenoxypropionate ACC inhibiting SEQ ID NO : 6 , or a Tryptophan to Cysteine amino acid herbicide , an cyclohexanedione ACC inhibiting herbicide or substitution at an amino acid position 1999 (W1999C ; SEQ an phenylpyrazolin ACC inhibiting herbicide. For example , ID NO : 7 ) aligning with the amino acid sequence of SEQ ID the ACC inhibitor herbicide tolerant sorghum plant or plant NO : 6 and a Tryptophan to Serine amino acid substitution at parts are tolerant or resistant when the AAC inhibiting an amino acid position 2027 W2027S ; SEQ ID NO : 10 ) herbicide is applied individually , or in a herbicide combi aligning with the amino acid sequence of SEQ ID NO : 6 , or nation , at a level that inhibits growth of a wild type sorghum a Tryptophan to Serine amino acid substitution at an amino plant. For example , the ACC inhibiting herbicide is clodi acid position 1999 (W1999S ; SEQ ID NO : 8 ) aligning with nafop -propargyl (CAS RN 105512 -06 - 9 ): cyhalofop -butyl the amino acid sequence of SEQ ID NO : 6 and an alanine to (CAS RN 122008 -85 - 9 ) ; diclofop -methyl (CAS RN 51338 valine amino acid substitution at an amino acid position 27 - 3 ); fenoxaprop - p - ethyl (CAS RN 71283 -80 - 2 ); fluazi 2004 (A2004V ; SEQ ID NO : 9 ) aligning with the amino acid fop - P - butyl (CAS RN 79241 - 46 -6 ) ; quizalofop - p - ethyl sequence of SEQ ID NO : 6 , or a Tryptophan to Serine amino (CAS RN 100646 -51 - 3 ) ; quizalofop - p (CAS RN 94051- 08 acid substitution at an amino acid position 1999 (W1999S ; 8 ) ; haloxyfop (CAS RN 69806 - 34 - 4 ) ; haloxyfop - ethoxy SEQ ID NO : 8 ) aligning with the amino acid sequence of ethyl (CAS RN 87237 - 48 - 7 ) ; haloxyfop - etotyl (CAS RN SEQ ID NO : 6 and a Tryptophan to Serine amino acid 87237 -48 - 7 ) ; haloxyfop - R -methyl (CAS RN 72619 - 32 -0 ); substitution at an amino acid position 2027 ( W2027S ; SEQ metamifop (CAS RN 256412 -89 -2 ); propaquizafop (CAS ID NO : 10 ) aligning with the amino acid sequence of SEQ RN 111479 - 05 - 1 ); alloxydim (CAS RN 55634 - 91 - 8 ) ; ID NO : 6 , or an alanine to valine amino acid substitution at butroxydim (CAS RN 138164 - 12 - 2 ) ; cycloxydim (CAS RN an amino acid position 2004 (A2004V ; SEQ ID NO : 9 ) 101205 -02 - 1 ) ; clethodim (CAS RN 99129 - 21 - 2 ) ; profoxy aligning with the amino acid sequence of SEQ ID NO : 6 and dim (CAS RN 139001 - 49 - 3 ) ; sethoxydim (CAS RN 74051 a Tryptophan to Serine amino acid substitution at an amino 80 - 2 ) ; tepraloxydim (CAS RN 149979 -41 - 9 ) ; tralkoxydim acid position 2027 (W2027S ; SEQ ID NO : 10 ) aligning with (CAS RN 87820 - 88 - 0 ); or pinoxaden (CAS RN 243973 - 20 the amino acid sequence of SEO ID NO : 6 , or a Tryptophan 8 ) . to Cysteine amino acid substitution at an amino acid position [0024 ] In some embodiments , the ACC inhibitor herbicide 1999 (W1999C ; SEQ ID NO : 7 ) aligning with the amino tolerant sorghum plants or plant that are homozygous or acid sequence of SEQ ID NO : 6 and a Tryptophan to Serine heterozygous for one or more of mutation of the ACC gene amino acid substitution at an amino acid position 1999 provided in the present disclosure. Alternatively , the ACC (W1999S ) aligning with the amino acid sequence of SEO ID inhibitor herbicide tolerant sorghum plants or plant parts NO : 6 and an alanine to valine amino acid substitution at an comprise one or more mutations of the ACC gene disclosed amino acid position 2004 (A2004V ; SEQ ID NO : 9 ) aligning in the present disclosure in homozygous or heterozygous with the amino acid sequence of SEQ ID NO : 6 , or a combinations. Tryptophan to Cysteine amino acid substitution at an amino [0025 ] In another embodiment, the invention provides for acid position 1999 (W1999C ; SEQ ID NO : 7 ) aligning with one or more Acetyl- CoA carboxylase (ACC ) inhibiting the amino acid sequence of SEQ ID NO : 6 and a Tryptophan herbicide capable of being used for controlling unwanted to Serine amino acid substitution at an amino acid position vegetation in one or more sorghum growing areas , wherein 1999 (W1999S ; SEQ ID NO : 8 ) aligning with the amino the sorghum plants in the growing area comprise one or acid sequence of SEQ ID NO : 6 and a Tryptophan to Serine more ACC inhibitor herbicide tolerant sorghum plants pro amino acid substitution at an amino acid position 2027 vided in the present disclosure. For example , the Acetyl (W2027S ; SEQ ID NO : 10 ) aligning with the amino acid COA carboxylase (ACC ) inhibiting herbicide include ACC sequence of SEQ ID NO : 6 , or a Tryptophan to Cysteine inhibiting herbicide is clodinafop -propargyl (CAS RN amino acid substitution at an amino acid position 1999 105512 - 06 - 9 ) : cyhalofop -butyl (CAS RN 122008 - 85 - 9 ) ; (W1999C ; SED ID NO : 7 ) aligning with the amino acid diclofop -methyl (CAS RN 51338 - 27 - 3 ) ; fenoxaprop - p - ethyl sequence of SEQ ID NO : 6 and an alanine to valine amino (CAS RN 71283 - 80 - 2 ) ; fluazifop - P -butyl ( CAS RN 79241 acid substitution at an amino acid position 2004 (A2004V ; 46 -6 ); quizalofop - p -ethyl (CAS RN 100646 -51 - 3 ); quizalo US 2018 /0346920 A1 Dec . 6 , 2018

fop - p (CAS RN 94051 - 08 - 8 ) ; haloxyfop (CAS RN 69806 protein amino acid sequence , b ) selecting a cell, plant or 34 - 4 ) ; haloxyfop - ethoxyethyl (CAS RN 87237 - 48 - 7 ) ; plant part that expresses the mutated AAC protein and grows haloxyfop -etotyl (CAS RN 87237 -48 - 7) ; haloxyfop -R in the presence of up to 200 uM of an ACC inhibitor methyl ( CAS RN 72619 -32 - 0 ); metamifop ( CAS RN herbicide , and c ) regenerating plant shoots from the selected 256412 - 89 - 2 ); propaquizafop (CAS RN 111479 -05 - 1 ); cell , plant or plant part in the presence of an ACC inhibitor alloxydim (CAS RN 55634 -91 -8 ); butroxydim (CAS RN herbicide . In any of the methods of the invention , the 138164 - 12 - 2 ); cycloxydim (CAS RN 101205 -02 - 1 ) ; transgene nucleotide sequence is derived from any source . clethodim (CAS RN 99129 - 21- 2 ); profoxydim (CAS RN [0029 ] In any of the method of the invention , the plant 139001 - 49 - 3 ) ; sethoxydim (CAS RN 74051 - 80 - 2 ) ; tepral cells are transformed with any method known in the art. For oxydim (CAS RN 149979 - 41 - 9 ) ; tralkoxydim (CAS RN example , the plant cell is transformed through PEG medi 87820 -88 -0 ); or pinoxaden (CAS RN 243973 - 20 - 8 ) . ated protoplast transformation , protoplast electroporation , [ 0026 ] In another embodiment, the invention provides for biolistics, or agrobacterium mediated transformation . In methods for introducing creating an Acetyl- CoA carboxy addition , the biolistic transformation is biolistic using lase (ACC ) inhibitor herbicide tolerant sorghum plant or embryogenic plant part having one or more mutations in the Acetyl - CoA [ 0030 ] In any of the method of the invention which Carboxylase ( ACC ) gene comprising the steps of: exposing comprise a step of regenerating plant shoots from aselected a sorghum plant or plant part to about 1 uM - 200 uM of an cells , the plants shoots are regenerated at an efficiency of ACC inhibitor herbicide , selecting a cell , plant or plant part 25 % or greater , 30 % or greater , 35 % or greater , 40 % or which grows in the presence of up to 200 uM of an ACC greater , 45 % or greater , 50 % or greater or 60 % greater . In inhibitor herbicide , and regenerating plant shoots from the addition , in any of the method of the invention which selected cell , plant or plant part in the presence of an ACC comprise a step of regerating plant shoots from aselected inhibitor herbicide . In these methods , the Acetyl- CoA car cells , the efficiency of regenerating a sorghum plant is 25 % boxylase (ACC ) inhibiting herbicide is clodinafop - propar or greater, 30 % or greater , 35 % or greater , 40 % or greater , gyl (CAS RN 105512 - 06 - 9 ): cyhalofop -butyl (CAS RN 45 % or greater , 50 % or greater or 60 % greater . 122008 -85 - 9 ) ; diclofop -methyl (CAS RN 51338 - 27 - 3 ) ; [0031 ] In any of the methods of the invention , the ACC fenoxaprop - p - ethyl (CAS RN 71283 - 80 - 2 ) ; fluazifop - P -bu inhibitor herbicide tolerant sorghum plant or plant part tyl (CAS RN 79241 -46 - 6 ) ; quizalofop - p - ethyl (CAS RN comprises one or more of the mutation of the Acetyl- CoA 100646 - 51- 3 ); quizalofop -p (CAS RN 94051 -08 -8 ); haloxy carboxylase (ACC ) gene disclosed herein . fop (CAS RN 69806 - 34 - 4 ) ; haloxyfop - ethoxyethyl (CAS [0032 ] In another embodiment, the invention provides for RN 87237 -48 - 7 ) ; haloxyfop - etotyl (CAS RN 87237 - 48 - 7 ) ; methods of producing ACC inhibitor herbicide tolerant haloxyfop - R -methyl (CAS RN 72619 - 32 - 0 ) ; metamifop sorghum plant progeny comprising the steps of a ) crossing (CAS RN 256412 - 89 - 2 ) ; propaquizafop (CAS RN 111479 a first ACC inhibitor herbicide tolerant sorghum plant dis 05 - 1 ) ; alloxydim (CAS RN 55634 - 91- 8 ) ; butroxydim (CAS closed herein with a second sorghum plant having a different RN 138164 - 12 - 2 ) ; cycloxydim (CAS RN 101205 - 02 - 1 ) ; genetic background , and b ) selecting a progeny plant result clethodim (CAS RN 99129 - 21 - 2 ) ; profoxydim (CAS RN ing from the crossing wherein the progeny comprises the 139001- 49 - 3 ) ; sethoxydim (CAS RN 74051 - 80 - 2 ) ; tepral mutation in the ACC gene of the first ACC inhibitor herbi oxydim (CAS RN 149979 - 41 - 9 ) ; tralkoxydim (CAS RN cide tolerant sorghum plant. For example , the crossing step 87820 -88 -0 ); or pinoxaden (CAS RN 243973 - 20 - 8 ) . comprises transferring pollen from the first ACC inhibitor [ 0027 ] The invention also provides for methods of creat herbicide tolerant sorghum plant to a wild -type sorghum ing an Acetyl- CoA carboxylase ( ACC ) herbicide tolerant plant and said crossing results in a population of progeny sorghum plant or plant part having one or more mutations in plants comprising the mutation of the first ACC inhibitor the Acetyl- CoA Carboxylase (ACC ) gene, comprising the herbicide tolerant sorghum plant. Alternatively , the crossing steps of a ) mutating the endogenous nucleotide sequence step comprises planting sterile female sorghum lines grown encoding the ACC protein by inserting, deleting , modifying and pollen shedding sorphum lines in isolated fields , or replacing one or more nucleotides within the genome of wherein one or both of the sorghum lines are ACC inhibitor living sorghum tissue using an engineered nuclease that herbicide tolerant sorghum plant disclosed herein , wherein creates site - specific double - strand breaks (DSBs ) ata the crossing results in hybrid seed comprising the mutation desired location in the genome, b ) selecting a cell , plant or of the first ACC inhibitor herbicide tolerant sorghum plant . plant part comprising the mutation and wherein the plant or In any of these methods, the progeny comprise one or more plant part grows in the presence of up to 200 uM of an ACC mutation of the Acetyl- CoA carboxylase (ACC ) gene dis inhibitor herbicide , and c ) regenerating plant shoots from the closed herein . selected cell , plant or plant part in the presence of an ACC [0033 ] The invention also provides for methods of devel inhibitor herbicide . For example , in any of these methods, oping a population of Acetyl- CoA carboxylase (ACC ) the endogenous nucleotide sequence encoding the ACC inhibitor herbicide tolerant sorghum plants comprising the protein is mutated using Meganuclease , Zinc -Fingure steps of a ) screening a population of sorghum plants to Nuclease , TALEN , or CRISPR technologies . identify a plant comprising one ormore of the mutations of [0028 ] In another embodiment, the invention provides for the ACC gene disclosed herein , and b ) propagating the methods of creating an Acetyl- CoA carboxylase (ACC ) identified sorghum plants comprising a mutation in the ACC herbicide tolerant sorghum plant or plant part having one or gene nucleotide sequence to develop a population of ACC more mutations in the Acetyl- CoA Carboxylase ( ACC ) inhibitor herbicide tolerant sorghum plants . For example , the gene , comprising the steps of a ) transforming a plant cell the screening step comprises using DNA markers to identify with one or more expression vectors , wherein the expression the ACC inhibitor herbicide tolerant sorghum plantor plant vector comprises a transgene nucleotide sequence , wherein part . For example , the screening step comprises applying the transgene nucleotide sequence encodes a mutated ACC ACC inhibiting herbicides on the population of sorghum US 2018 /0346920 A1 Dec. 6 , 2018

plants . In any of these methods, the ACC inhibiting herbi - 51338 -27 -3 ) ; fenoxaprop -p - ethyl (CAS RN 71283 -80 -2 ) ; cides is applied using a spray carrier , wherein the herbicide fluazifop - P -butyl (CAS RN 79241 -46 - 6 ) ; quizalofop - p is applied at two to four times the recommended rate of ethyl ( CAS RN 100646 -51 - 3 ) ; quizalofop - p (CAS RN herbicide application per area of land or is applied at two to 94051- 08 - 8 ) ; haloxyfop (CAS RN 69806 - 34 -4 ); haloxyfop four times the herbicide concentration per volume of carrier. ethoxyethyl (CAS RN 87237 -48 - 7 ) ; haloxyfop - etotyl (CAS In addition , in any of these methods further comprise the RN 87237 -48 -7 ); haloxyfop - R -methyl (CAS RN 72619 -32 step of selecting healthy plants 14 days after herbicide 0 ) ; metamifop ( CAS RN 256412 -89 - 2 ) ; propaquizafop application to identify herbicide tolerant sorghum plants . (CAS RN 111479 -05 - 1 ) ; alloxydim (CAS RN 55634 - 91 - 8 ) ; [0034 ] In any of the methods of the invention which butroxydim (CAS RN 138164 - 12 - 2 ) ; cycloxydim (CAS RN involve applying the ACC inhibiting herbicide, said herbi 101205 - 02 - 1 ) ; clethodim (CAS RN 99129 - 21 - 2 ); profoxy cide is applied to one of the following : a segregating dim (CAS RN 139001 - 49 -3 ); sethoxydim (CAS RN 74051 population of inbred lines in the field , greenhouse or growth 80 - 2 ) ; tepraloxydim (CAS RN 149979 -41 - 9 ) ; tralkoxydim chamber, wherein the resulting inbred lines are tolerant to (CAS RN 87820 - 88 - 0 ) ; or pinoxaden (CAS RN 243973 - 20 ACC inhibiting herbicides and eliminate wild - type sorghum 8 ). plants ; a field of sterile female A - lines , restorer male [0037 ] In particular, the ACC inhibiting herbicide is R - Lines , or both inbred lines in a breeding nursery where Quizalofop - p - ethyl, Clethodim or a mixture thereof , manual pollination or crossing is conducted for the produc wherein the dose of Quizalofop - p - ethyl is equivalent to 6 . 3 tion of hybrid seed , wherein the resulting hybrid seed is g a . i/ ha and the dose of Clethodim is equivalent to 12 . 5 g tolerant to ACC inhibiting herbicides and eliminate wild a . i/ ha and the AAC inhibiting herbicide is applied to one of type sorghum plants ; a field or greenhouse containing both the following : a field or greenhouse containing sterile female sterile female parent A - lines and restorer male parent parent A - lines , restorer male parent R - Lines , or both inbred R -Lines , or both parent inbred lines , wherein one or both parent lines , wherein one or both parents are tolerant to ACC parents are tolerant to ACC inhibiting herbicides, for pro inhibiting herbicides , for production of hybrid seed that is duction of hybrid seed that is tolerant to ACC inhibiting tolerant to ACC inhibiting herbicides ; or a grain production herbicides and to eliminate wild type sorghum plants ; or a field in which hybrid ( F1) seed has been planted , wherein the grain production field in which hybrid (F1 ) seed was hybrid seed is tolerant to ACC inhibiting herbicides for the planted , wherein the seed is tolerant to ACC inhibiting purpose of controlling weeds and producing sorghum grain herbicides to eliminate wild type sorghum plants . or forage . [0035 ] In any of these methods, the ACC inhibiting her [0038 ] In another embodiment, the invention provides for bicides that is applied is clodinafop - propargyl (CAS RN methods for controlling unwanted vegetation in a sorghum 105512 -06 - 9 ) : cyhalofop -butyl (CAS RN 122008 - 85 - 9 ) ; plant growing area comprising an ACC inhibitor herbicide diclofop -methyl (CAS RN 51338 -27 -3 ) ; fenoxaprop - p -ethyl tolerant sorghum plant of any one of claims 1 - 8 with one or (CAS RN 71283 - 80 - 2 ) ; fluazifop - P -butyl (CAS RN 79241 more ACC inhibitor herbicide ( s ) , wherein the AAC inhibitor 46 - 6 ); quizalofop -p - ethyl (CAS RN 100646 - 51 -3 ) ; quizalo herbicide is applied alone or in combination with one or fop - p (CAS RN 94051 -08 - 8 ) ; haloxyfop (CAS RN 69806 more non - ACC inhibitor herbicide . In these methods , the 34 - 4 ) ; haloxyfop - ethoxyethyl (CAS RN 87237 - 48 - 7 ) ; AAC inhibitor herbicide and the non -ACC inhibitor herbi haloxyfop - etotyl (CAS RN 87237 - 48 - 7 ) , haloxyfop - R cide are applied jointly or simultaneously . Alternatively , the methyl (CAS RN 72619 -32 -0 ); metamifop (CAS RN AAC inhibitor herbicide and the non - ACC inhibitor herbi 256412 -89 - 2 ); propaquizafop (CAS RN 111479- 05 - 1 ) ; cide are applied at different times . In addition , in these alloxydim (CAS RN 55634 - 91 -8 ); butroxydim (CAS RN methods the AAC inhibitor herbicide and the non - ACC 138164 - 12 - 2 ) , cycloxydim ( CAS RN 101205 - 02 - 1 ) ; inhibitor herbicide are applied sequentially , in pre - emer clethodim ( CAS RN 99129 - 21- 2 ) ; profoxydim (CAS RN gence applications followed by post -emergence applica 139001 - 49 - 3 ) ; sethoxydim (CAS RN 74051 - 80 - 2 ) ; tepral tions , or in early post- emergence applications followed by oxydim ( CAS RN 149979 -41 - 9 ) ; tralkoxydim (CAS RN medium or late post -emergence applications. 87820 -88 -0 ); or pinoxaden (CAS RN 243973 - 20 - 8 ) . In [0039 ] In any of these methods, the ACC herbicide ( s ) is particular, the ACC inhibiting herbicide is Quizalofop - p clodinafop -propargyl (CAS RN 105512 -06 - 9 ): cyhalofop ethyl or Clethodim or a mixture thereof, wherein the dose of butyl ( CAS RN 122008 -85 - 9 ) ; diclofop -methyl (CAS RN Quizalofop - p -ethyl is equivalent to 6 . 3 g a .i /ha and the dose 51338 - 27 - 3 ); fenoxaprop - p - ethyl (CAS RN 71283 -80 - 2 ); of Clethodim is equivalent to 12 . 5 g a . i/ ha and the herbicide fluazifop - P - butyl (CAS RN 79241 - 46 - 6 ) ; quizalofop - p is applied to a segregating population of inbred lines in the ethyl (CAS RN 100646 -51 - 3 ) ; quizalofop -p (CAS RN field , greenhouse or growth chamber to create new ACC 94051 -08 - 8 ) ; haloxyfop (CAS RN 69806 - 34 - 4 ) ; haloxyfop inhibiting herbicide tolerant inbred lines. ethoxyethyl (CAS RN 87237 -48 - 7 ) ; haloxyfop - etotyl ( CAS [0036 ] In another embodiment, the invention provides for RN 87237 -48 - 7 ) ; haloxyfop - R -methyl (CAS RN 72619 - 32 a method of using an ACC inhibitor herbicide tolerant 0 ) ; metamifop (CAS RN 256412 -89 - 2 ) ; propaquizafop sorghum plant or plant part of any one of claims 1 - 8 , for the (CAS RN 111479 -05 - 1 ); alloxydim (CAS RN 55634 -91 -8 ); elimination of unwanted vegetation or for the production of butroxydim (CAS RN 138164 - 12 - 2 ) ; cycloxydim (CAS RN seed or grain , comprising applying a mixture comprising 101205 -02 - 1) ; clethodim (CAS RN 99129 -21 - 2 ) ; profoxy one or more AAC inhibiting herbicides and a spray carrier, dim (CAS RN 139001 - 49 - 3 ) ; sethoxydim (CAS RN 74051 wherein the mixture is applied at a recommended rate of 80 - 2 ) ; tepraloxydim (CAS RN 149979 -41 - 9 ); tralkoxydim herbicide per area of land or concentration per volume of (CAS RN 87820 - 88 - 0 ) ; or pinoxaden (CAS RN 243973 - 20 carrier for the control of weedy grasses in other tolerant 8 ) . crops. In any of these methods, the AAC inhibiting herbicide [0040 ] In any of the methods of the invention which is clodinafop -propargyl (CAS RN 105512 - 06 - 9 ) : cyhalofop comprise applying a non -AAC inhibiting herbicide , the butyl (CAS RN 122008 - 85 - 9 ) ; diclofop -methyl (CAS RN non -ACC inhibiting herbicide is one of the following an US 2018 /0346920 A1 Dec . 6 , 2018 inhibitor of lipid synthesis such as aryloxyphenoxypropi- bicide Assure II at 10 oz /acre (2x ) and photographed 2 onate , a cyclohexanodeione , a benzofurane , a chloro - car weeks after herbicide application . Arrow indicates tissue bonic acid , a phosphorodithioate, a phyenylpyrazolin or a culture derived BTX430 control plants . thiocarbamate ; an inhibitor of photosynthesis at photosys [0048 ] FIG . 8 provides screening results of herbicide tem II such as phenyl- carbamate , a pyridazinone, a triazine , resistance in young (2 weeks old ) tissue culture derived F . a triazinone , a triazolinone , an uracil , an amide , an urea , a sorghum plants ( FPS ) containing W2027S mutation . Plants benzothiadiazinone, a nitrile or a phenyl- pyridine ; an inhibi were sprayed with low rate of quizalofop herbicide Assure tor of photosynthesis at photosystem I such as bipyridylium ; II at 10 ozlacre (2x ) and photographed 2 weeks after an inhibitor of protoporphyrinogen oxidase such as diphe herbicide application . Arrow indicates tissue culture derived nylether, a N -phenylphthalimide , an oxadiazole, an oxyzo BTX430 control plants . Screening results of herbicide resis lidinedione, a phenylpyrazole , a pyrimidindione , or a thia tance in young ( 2 weeks old ) tissue culture derived F . diazol; an inhibitor of carotenoid biosynthesis such as sorghum plants (FPS ) containing W2027S mutation . Plants pyridazinone, a pyridinecarboxamide , an isoxazolidinone , were sprayed with low rate of quizalofop herbicide Assure or a triazole ; an inhibitor of 4 -hydroxyphenyl - pyruvate - II at 10 oz /acre (2x ) and photographed 2 weeks after callistemone such as isoxazole, a pyrazole , or a triketone ; an herbicide application . Arrow indicates tissue culture derived inhibitor of EPSP synthase such as glycine ; an inhibitor of BTX430 control plants . glutamine synthesis such as phosphinic acid ; an inhibitor of 10049 ] FIG . 9 demonstrates herbicide resistance in dihydropteroate synthase such as carbamate ; an inhibitors of matured ( 6 - 8 weeks old ) tissue culture derived F , sorghum microtubule assembly such as benzamide , a benzoic acid , a plants (BTX430 ) containing W1999C mutation . Plants were dinitroaniline , a phosphoroamidate or a pyridine ; an inhibi sprayed with high field rate application of Assure II at 8 tor of cell division such as acetamide, a chloroacetamide, or oz /acre (1x ) , 16 oz / acre ( 2x ) and 32 oz /acre (4x ) and an oxyacetamide ; an inhibitor of cell wall synthesis such as photographed 3 weeks after herbicide application . nitrile or a triazolocarboxamide ; or an inhibitor of auxin [0050 ] FIG . 10 demonstrates herbicide resistance in Fi transport such as a phthalamate or a semicarbazone. heterozygous sorghum plants (BTX430 ) containing W1999C mutation . Plants were sprayed at 2x rate of Assure BRIEF DESCRIPTION OF DRAWING II ( 16 oz / acre ) with spray volume of 15 gallons/ acre . Plants [ 0041] FIG . 1 provides FOP herbicide resistant calli (FP were photographed 10 days after herbicide application . 13 ) growing on tissue culture medium containing 1 , 1 . 5 , 2 , [0051 ] FIG . 11 provides Fi plant of BTX430 herbicide 5 , 10 uM of Quizalofop -p - ethyl and 5 uM of Assure II resistant plants (W1999C ) with good seed set under green herbicide and house conditions . [ 0042 ] FIG . 2 provides FOP resistant calli (FP - 7 , FP - 8 , [0052 ] FIG . 12 demonstrates KASP assay developed for FP - 9 , FP - 10 , FP - 13, FP - 15) surviving on 100 uM of detecting SNP at W1999C codon position of ACC gene . Quizalofop - p - ethyl ( chemical A . I. ) . Three clusters separate homozygous mutants ( TGC ) , het [ 0043] FIG . 3 demonstrates regeneration and rooting of erozygous mutants ( TG ( G / C ) and wild types allele ( TGG ) . FOP resistant ( FP - 13 ) and control BTX430 calli on media [0053 ] FIG . 13A -FIG . 13B demonstrates complete herbi containing 0 .0 and 1. 0 uM Quizalofop - p - ethyl ( chemical cide resistance in homozygous F2 BTX430 plants with A .I . ) . W1999C mutation . Picture shows before (A ) and 14 days [0044 ] FIG . 4 provides DNA and protein sequences of the after ( B ) herbicide application at 2x rate ( 16 oz /acre ) . carboxyl transferase region of sorghum ACC gene . SEQ ID 100541. FIG . 14A - FIG . 14B . Complete herbicide resistance NOS : 1 and 6 are DNA and protein sequences of wild type in homozygous F , BTX430 plants with W1999C mutation . sorghum (BTX430 ) . SEQ ID NOS: 2 and 7 , 3 and 8 , 4 and Pictures shows before ( A ) and 14 days after ( B ) herbicide 9 , 5 and 10 represents the DNA and protein sequence of application at 4x rate ( 32 oz / acre ) . mutated ACC1, ACC2, ACC3 and ACC4 , respectively with mutation at W1999C , W1999S , A2004V and W2027S codon DETAILED DESCRIPTION position ( W = Tryptophan , C = Cysteine , S = Serine , A = Alanine , V = Valine ). [0055 ] As used herein , the term “ variety ” and “ cultivar ” [0045 ] FIG . 5 provides a comparison of the four amino refers to plants that are defined by the expression of the acid mutations at W1999C , W1999S , A2004V and W2027S characteristics resulting from a given genotype or combina in the CT domain of sorghum ACC gene found to be tion of genotypes , distinguished from any other plant group associated with ACC herbicide resistance in sorghum line ing by the expression of at least one of the characteristics BTX430 -CHR - ACCs. ( W = Tryptophan , C = Cysteine , and considered as a unit with regard to its suitability for S = Serine, A = Alanine , V = Valine ). being propagation unchanged . [ 0046 ] FIG . 6 provides screening results of herbicide [0056 ] As used herein , the term “ hybrid ” refers to the resistance in young ( 2 weeks old ) tissue culture derived F . offspring or progeny of genetically dissimilar plant parents sorghum plants (BTX430 ) containing W1999C mutation . or stock produced as the result of controlled cross - pollina Plants were sprayed with low rate of quizalofop herbicide tion , or by commercial hybrid seed production in which Assure II at 2 .5 oz / acre (0 .5x ), 5 oz /acre ( 1x ) and 10 oz /acre male and female lines are planted near to each other. ( 2x ) and photographed 2 weeks after herbicide application . [0057 ] As used herein , the term “ progeny ” refers to gen Arrow indicates tissue culture derived BTX430 control erations or offspring of a plant. plants . [ 0058 ] As used herein , the term " derivative ” of an herbi [0047 ] FIG . 7 provides screening results of herbicide cide resistant plant includes both the progeny of that herbi resistance in young ( 2 weeks old ) tissue culture derived F . cide resistant plant, as well as any mutant, recombinant , or sorghum plants (FP11 and FP12 ) containing W1999S muta genetically engineered derivative of that plant, whether of tion . Plants were sprayed with low rate of quizalofop her the same species or a different species , where the herbicide US 2018 /0346920 A1 Dec . 6 , 2018 resistant characteristic (s ) of the original herbicide resistant eign genetic material into a line of breeding stock . For plant has been transferred to the derivative plant. example , the present invention provides for sorghum crop [ 0059 ] As used herein , the term “ plant tissue” includes plants introgressed with a mutant ACC gene for herbicide differentiated and undifferentiated tissues of plants including tolerance by crossing two plant generations. those present in roots, shoots , leaves, pollen , seeds and 10070 ] As used herein , the term “herbicide tolerant” or tumors , as well as cells in culture ( e . g . , single cells , proto “ herbicide tolerance” refers to an improved capacity of a plasts , embryos, callus , etc . ) . Plant tissue may be in planta , particular plant to withstand the various degrees of herbi in organ culture , tissue culture , or cell culture . cidally induced injury that typically are lethal to wild - type [ 0060 ] As used herein , the term “ plant part” as used herein plants of the same genotype at the sameherbicidal dose. The refers to a plant structure or a plant tissue, for example , term " herbicide resistant” or “ herbicide resistance ” refers to pollen , an ovule , a tissue, a pod , a seed , a leaves, panicles, the inherited ability of a plant to survive and reproduce roots , caryopsis , stem and a cell. In some embodiments of following exposure to a dose of herbicide normally lethal to the present invention transgenic plants are crop plants . the wild type . In a plant , resistance may be naturally [0061 ] As used herein , the term “ caryopsis ” as used herein occurring or induced by such techniques as genetic engi refers to a dry , single carpel and indehiscent fruit in which neering or selection of variants produced by tissue culture or the ovary wall is united with the seed coat, typically of grass mutagenesis . As used herein unless otherwise indicated , species . The caryopsis is commonly referred to as grain or herbicide “ resistance” is heritable and allows a plant to grow seed , with the use often dependent upon the final use . and reproduce in the presence of a typical herbicidally [0062 ] As used herein , the terms " crop " and " crop plant" effective treatment by a herbicide for a given plant, as are used in their broadest sense . The term includes, but is not suggested by the current edition of The Herbicide Handbook limited to , any species of plant edible by humans or used as as of the filing of the subject disclosure . As is recognized by a feed for animal or fish or marine animal, or consumed by those skilled in the art , a plant may still be considered humans, or used by humans , or viewed by humans, or any “ resistant” even though some degree of plant injury from plant used in industry , commerce , or education . herbicidal exposure is apparent. As used herein , the term [0063 ] As used herein , the terms “ F -generation ” and “ filial " tolerance ” or “ tolerant” includes “ resistance ” or “ resistant" generation ” refers to any of the consecutive generations of plants as defined herein , as well an improved capacity of a plants , cells , tissues or organisms after a biparental cross. particular plant to withstand the various degrees of herbi The generation resulting from a mating of the a biparental cidally induced injury that typically are lethyl in wild - type cross ( i . e . two parents ) is the first filial generation (desig plants of the same genotype at the same herbicidal dose . nated as “ F?” and “ F?” ) in reference to a seed and it' s plant, [ 0071 ] As used herein , the term “ wild -type ” when made in while that resulting from crossing of F individual is the reference to a gene refers to a functional gene common second filial generation (designated as " F , ” or “ F , ” ) in throughout a plant population . A functional wild - type gene reference to a seed and it' s plant. For example , an F2 seed is that which is most frequently observed in a population and and a resulting plant are produced by self- pollination or is thus arbitrarily designated the “ normal” or “wild -type ” cross -pollination of F1, while later F generations are pro form of the gene. duced from self -pollination or cross -pollination of the 10072 ] As used herein , the terms “ modified ” or “ mutant " immediate prior generation . or “ functional mutant” when made in reference to a gene or [ 0064 ] As used herein , the term “ germplasm ” refers to any to a gene product refers, respectively, to a gene or to a gene genetic material of plants that contain functional units of product which displays modifications in sequence and /or heredity . functional properties ( i . e . , altered characteristics ) when [0065 ] As used herein , the term “ elite germplasm ” in compared with the wild - type gene or gene product. Thus, the reference to a plant refers to hereditary material of proven terms "modified ” and “ mutant " when used in reference to a genetic superiority . nucleotide sequence refer to an nucleic acid sequence that [0066 ]. As used herein , the term “ elite plant, ” refers to any differs by one or more nucleotides from another , usually plant that has resulted from breeding and selection for related nucleotide acid sequence and the term " functional superior agronomic performance . mutant” when used in reference to a polypeptide encodes by 10067] As used herein , the term “ trait ” refers to an observ said “ modified ” or “ mutant” nucleic acid refers to the able and /measurable characteristic of an organism . For protein or polypeptide that retains activity . In the present example , the present invention describes plants that are application , the ACC mutant protein , “ or functionalmutant " resistant to FOP and DIM herbicides , whereby that resis thereof is an ACC gene that retains its native activity to tance is a trait . create essential amino acids . Additionally, a “ modified ” [0068 ] As used herein , the terms “marker ” and “ DNA nucleotide sequence is interpreted as that found in the marker " and " molecular marker" in reference to a “ select- degenerate genetic code as known by those skilled in the art . able marker ” refers to a physiological or morphological trait For example , the genetic code is degenerate as there are that may be determined as a marker for its own selection or instances in which different codons specify the same amino for selection of other traits closely linked to that marker. For acid ; a genetic code in which some amino acids may each be example , such a marker could be a gene or trait that encoded by more than one codon . It is contemplated that the associates with herbicide tolerance including, but not limited present invention may comprise such degeneracy ( e . g . , to , simple sequence repeat (SSR ), single nucleotide poly wherein a sorghum hybrid comprises an ACC gene that is at morphism (SNP ) , genetic insertions and / or deletions and the least 70 % identical or homologous , at least 80 % identical or like . homologous , at least 85 % identical or homologous , at least [0069 ] As used herein , the term “ introgress ” and “ intro 90 % identical or homologous, at least 95 % identical or gressing ” and “ introgression ” refers to conventional ( i. e . homologous, at least 97 % identicalor homologous, or at classic ) pollination breeding techniques to incorporate for - least 99 % identical or homologous to the nucleotide US 2018 /0346920 A1 Dec. 6 , 2018

sequence of SEQ ID NOS: 1 to 5 as found in , for example , 6 ; or a Tryptophan to Serine amino acid substitution at an the sorghum germplasm . Despite the differences in the amino acid position 1999 (SEQ ID NO : 8 ; W1999S ) align sequences the ACC genes of the invention , these ACC gene ing with SEQ ID NO : 6 ; or an Alanine to Valine amino acid sequence retain the ability to confer resistance and / or toler substitution at an amino acid position 2004 (SEQ ID NO : 9 ; ance to ACC inhibiting herbicides . The invention provides A2004V ) aligning with SEQ ID NO : 6 ; or a Tryptophan to for these AAV nucleotide sequence that are homologous or Serine amino acid substitution at an amino acid position exhibit sequence identity to SEQ ID NOS : 1 - 5 , and comprise 2027 ( SEQ ID NO : 10 ; W2027S ) aligning with SEQ ID NO : at least one of the mutations disclosed herein . 6 of sorghum Acetyl- CoA Carboxylase large subunit . [0073 ] As used herein , the term " heterologous ” when used [0081 ] The sorghum plant may also comprise all possible in reference to a gene or nucleic acid refers to a gene that has combination of the mutations and subsequent amino acid been manipulated in some way. substitutions shown in SEQ ID NOS : 2 , 3 , 4 , and 5 and SEQ [0074 ] As used herein , the term “ portion " or " functional ID Nos. 7 , 8 , 9 and 10 , respectively . fragment" when used in reference to a protein (as in “ a [0082 ] As such , the sorghum plantmay be tolerant to any fragment of a given protein ” , “ a protein fragment” , a “ por herbicide or combination of herbicides capable of inhibiting tion of a protein ” ) refers to fragments of that protein . The ACC enzyme activity , i. e . , the sorghum plant may be toler fragments may range in size from four amino acid residues ant to herbicides of the FOP family , such as , without to the entire amino sequence minus one amino acid . In the limitation , Cyhalofop -butyl (CAS RN 122008 - 85 - 9 ) ; Diclo present invention , the protein fragment is preferentially fop -methyl (CAS RN 51338 - 27 - 3 ) ; Fenoxaprop - P -ethyl functional such that the protein fragment confers resistance ( CAS RN 71283 -80 - 2 ); Fluazifop - P -butyl (CAS RN 79241 to inhibition to ACC herbicides to a given plant. 46 - 6 ); Quizalofop - P (CAS RN 76578 - 12 -6 ) ; Haloxyfop [ 0075 ] When used herein , the term “ transgenic ”” means (CAS RN 69806 - 34 - 4 ) ; Metamifop (CAS RN 256412 -89 that a gene — which can be of the same or a different 2 ) ; Propaquizafop (CAS RN 111479- 05 - 1 ) , or herbicides species has been introduced via an appropriate biological from the DIM family , such as, without limitation , Clethodim carrier, like Agrobacterium tumefaciens or by any other (CAS RN 99129 - 21 - 2 ) ; Sethoxydim (CAS RN 74051- 80 - 2 ) ; physical means, like protoplast transformation or particle Tepraloxydim (CAS RN 149979 - 41 -9 ); Tralkoxydim (CAS bombardment, into a plant and which gene is able to be RN 87820 -88 - 0 ), or herbicides from the DEN family , such expressed in the new host environment, namely the geneti as , without limitation , Pinoxaden (CAS RN 243973 - 20 - 8 ) . cally modified organism (GMO ) . [ 0083 ] The “ CAS RN ” stated in parentheses behind the 10076 ]. As used herein , the term " gene editing ” is a type of names corresponds to the chemical abstract service registry genetic engineering in which DNA is inserted , deleted , number ” , a customary reference number which allows the modified or replaced in the genome of a living organism . substances named to be classified unambiguously , since the The common methods for such editing use engineered “ CAS RN ’ distinguishes, inter alia , between isomers includ sequence - specific nucleases, or " molecular scissors” . These ing stereoisomers . nucleases create site - specific double -strand breaks (DSBs ) [0084 ] In a preferred embodiment, the sorghum plant of at desired locations in the genome. The induced double the invention which is resistant to herbicides belonging to strand breaks are repaired through nonhomologous end the groups FOP, DIM , or DEN herbicides is the sorghum line joining (NHEJ ) or homologous recombination (HR ) , result designated BTX430 - CHR -ACCI , BTX430 - CHR -ACC2 ing in targeted mutations (“ edits ” ) . and BTX430 -CHR - ACC4 which seed were deposited with [0077 ] In accordance to the before definition , the term the ATCC , Manassas , Va. , under Number PTA - 125106 , " non - transgenic ' means exactly the contrary , i . e . that no PTA - 125108 and PTA - 125107 respectively on May 9 , 2018 , introduction of the respective gene has occurred via an under the terms of the Budapest Treaty . The mutated appropriate biological carrier or by any other physical BTX430 - CHR - ACC1 plant, parts thereof and it seeds , com means . However, a mutated gene can be transferred through prise in the genome mutated ACC gene comprising at least pollination , either naturally or via a breeding process to one polynucleotide encoding a polypeptide having a Tryp produce another non - transgenic plant concerning this spe tophan to Cysteine amino acid substitution at an amino acid cific gene . position 1999 (SEQ ID NO : 7 ; W1999C ) aligning with SEQ [ 0078 ] An “ endogenous gene” means a gene of a plant ID NO : 6 of sorghum Acetyl- CoA Carboxylase large sub which has not been introduced into the plant by genetic unit . The mutated BTX430 - CHR - ACC2 plant, parts thereof engineering techniques. and it seeds, comprise in the genome mutated ACC gene [0079 ] The term “ sequence ” when used herein relates to comprising at least one polynucleotide encoding a polypep nucleotide sequence ( s) , polynucleotide (s ) , nucleic acid tide having a Tryptophan to Serine amino acid substitution sequence ( s ) , nucleic acid ( s ) , nucleic acid molecule , pep at an amino acid position 1999 (SEQ ID NO : 8 ; W1999S ) tides , polypeptides and proteins, depending on the context in aligning with SEQ ID NO : 6 of sorghum Acetyl- CoA which the term “ sequence ' is used . Carboxylase large subunit . The mutated BTX430 - CHR [0080 ] The sorghum plants in the current invention show ACC4 plant, parts thereof and it seeds , comprise in the improved resistance to herbicides , for example herbicides genome mutated ACC gene comprising at least one poly targeting the ACC enzyme , such as aryloxyphenoxypropi nucleotide encoding a polypeptide having a or a Tryptophan onates (FOP ) , cyclohexanediones (DIM ) and phenylpyrazo to Serine amino acid substitution at an amino acid position lins (DEN ) , as compared with wild - type sorghum plants. In 2027 (SEQ ID NO : 10 ;W2027S ) aligning with SEQ ID NO : particular, the sorghum plant (Sorghum bicolor ) of the 6 of sorghum Acetyl- CoA Carboxylase large subunit . present invention comprises in its genome at least one [0085 ] In a preferred embodiment, the herbicide -resistant polynucleotide encoding a polypeptide having a Tryptophan sorghum plant comprises the resistance traits of BTX430 to Cysteine amino acid substitution at an amino acid position CHR - ACC1, BTX430 -CHR -ACC2 and BTX430 -CHR 1999 (SEQ ID NO : 7 ; W1999C ) aligning with SEQ ID NO : ACC4 which seed were deposited with the ATCC , Manas US 2018 /0346920 A1 Dec . 6 , 2018 sas, Va. , under Number PTA - 125106 , PTA - 125108 and plants with both these attributes could be crossed using PTA - 125107 , and may be a plant as described in BTX430 - classical breeding techniques . In the pedigree method , supe CHR - ACCI, BTX430 - CHR - ACC2 and BTX430 - CHR rior plants are selfed and selected in successive generations . ACC4 which seed were deposited with the ATCC , a progeny In the succeeding generations , the heterozygous condition of BTX430 - CHR - ACC1, BTX430 -CHR - ACC2 and gives way to homogeneous lines as a result of self- pollina BTX430 - CHR -ACC4 which seed were deposited with the tion and selection . Typically , in the pedigree method , five or ATCC , Manassas, Va ., under Number PTA - 125106 , PTA more generations of selfing and selection are practiced ( e . g . , 125108 and PTA - 125107 , a mutant of BTX430 - CHR S1, S2, S3, S4, S5 , etc . ) . ACC1, BTX430 - CHR -ACC2 and BTX430 -CHR - ACC4 [0088 ] Backcrossing is used to improve a plant line . which seed were deposited with the ATCC , Manassas, Va ., Backcrossing transfers a specific desirable trait from one under Number PTA - 125106 , PTA - 125108 and PTA - 125107 , source to another that lacks the trait . This is accomplished and a progeny of BTX430 -CHR - ACC1, BTX430 -CHR by , for example , crossing a donor ( e . g . , BTX430 - CHR ACC2 and BTX430 - CHR - ACC4 which seed were deposited ACC1, BTX430 -CHR - ACC2 or BTX430 -CHR - ACC4 ) to with the ATCC ,Manassas , Va ., under Number PTA - 125106 , an inbred line (e . g. , an elite line as described herein ). The PTA - 125108 and PTA - 125107 mutant. Further provided is a progeny of this cross is then crossed back ( i . e . backcrossing ) sorghum seed , comprising in its genome at least one poly to the elite inbred line , followed by selection in the resultant nucleotide encoding a polypeptide having a Tryptophan to progeny for the desired trait ( e . g ., resistance to ACC herbi Cysteine amino acid substitution at an amino acid position cides ). Following at least one , and as many as five or more 1999 (SEQ ID NO : 7 ; W1999C ) aligning with SEQ ID NO : backcross generations with selection for the desired trait the 6 ; or a Tryptophan to Serine amino acid substitution at an progeny are typically heterozygous for the locus ( loci) amino acid position 1999 (SEQ ID NO : 8 ; W1999S ) align controlling the desired phenotype, but will be like the elite ing with SEQ ID NO : 6 ; or an Alanine to Valine amino acid parent for the other genetic traits . The last backcrossing then substitution at an amino acid position 2004 (SEQ ID NO : 9 ; is typically selfed in order to give a pure breeding progeny A2004V ) aligning with SEQ ID NO : 6 ; or a Tryptophan to for the gene being transferred . Serine amino acid substitution at an amino acid position [ 0089 ] In current hybrid sorghum breeding programs, new 2027 (SEQ ID NO : 10 ; W2027S ) aligning with SEQ ID NO : parent lines are developed to be either seed -parent lines 6 of sorghum Acetyl- CoA Carboxylase large subunit. The ( e .g ., Chromatin Proprietary B -lines B .791 , B . 1498 , and seed germinates and produces a plant having increased B .230 or pollen - parent lines ( e . g . , Chromatin Proprietary resistance to one or more herbicides of the FOP, DIM , or R -lines R .410 , R .M46 , and R . 373 depending on whether or DEN groups as compared with wild - type sorghum plants . In not they contain fertility restoring genes ; the seed - parent a preferred embodiment said seed is the seed deposited as lines do not have fertility restoring genes and are male PTA - 125106 , PTA - 125106 sterile in certain cytoplasms (also known as “ A ” line plants ) [ 0086 ] Sorghum plants are self -pollinating plants , but they and male - fertile in other cytoplasms (also known as “ B ” line can also be bred by cross- pollination . The development of plants) , whereas the pollen -parent lines are not male sterile sorghum hybrids requires the development of pollinator and do contain fertility restoring genes ( also known as “ R ” parents ( fertility restorers ) and seed parent inbreds using the line plants ). The seed -parent lines are typically created to be cytoplasmic male sterility - fertility restorer system , the cytoplasmically male sterile such that the anthers are mini crossing of seed parents and pollinator parents , and the mal to non -existant in these plants thereby requiring cross evaluation of the crosses. Pedigree breeding programs com pollination . The seed -parent lines will only produce seed , bine desirable traits ; in the present invention , the desirable and the cytoplasm is transmitted only through the egg . The trait being plant resistance to ACC herbicides . This trait is pollen for cross pollination is furnished through the pollen put into the breeding pool from one or more lines, such that parent lines that contain the genes necessary for complete new inbred lines can be created by crossing , followed by fertility restoration in the F , hybrid , and the cross combines selection of plants with the desired trait , followed by more with the male sterile seed parent to produce a high - yielding crossing, etc . New inbreds are crossed with other inbred single cross hybrid with good grain quality . lines (e . g ., elite plant lines like those described herein ). [ 0090 ] Typically , this cytoplasmic male sterility - fertility [ 0087 ] Pedigree breeding starts with the crossing of two restorer system is performed for the production of hybrid genotypes , in a preferred embodiment, the two genotypes seed by planting blocks of rows ofmale sterile (seed -parent ) might be BTX430 -CHR - ACC1, and an elite sorghum line plants and blocks of rows of fertility restorer (pollen -parent ) ( e . g . , Chromatin Proprietary lines such as R . 410 , R . 159 , plants , such that the seed -parent plants are wind pollinated R . 373 ) or BTX430 -CHR - ACC2 , and an elite sorghum line with pollen from the pollen -parent plant. This process pro ( e . g . , Chromatin Proprietary lines such as R .410 , R . 159 , duces a vigorous single -cross hybrid that is harvested and R . 373 ) or BTX430 -CHR - ACC4, and an elite sorghum line subsequently purchased or acquired and planted by the ( e . g . , Chromatin Proprietary lines such as R .410 , R . 159 , consumer. Male sterile , seed - parent plants can also be cre R . 373 ) . BTX430 - CHR -ACC1 , BTX430 -CHR - ACC2 and ated by genetically breeding recessive male -sterile nuclear BTX430 -CHR - ACC4 which seed were deposited with the genes into a particular population , however the cytoplasmic ATCC , Manassas, Va ., under Number PTA - 125106 , PTA male sterility - fertility restorer system is typically the system 125108 and PTA - 125107 on May 9, 2018 . If the original two used for breeding hybrid sorghum . Sleper and Poehlman parents do not provide all of the desired characteristics, then 2006 , Breeding Field Crops, Fifth Ed . , Blackwell Publishing other sources can be included in the breeding population . provides a good review of current sorghum breeding pro For example, if a hybrid is desired such that both ACC cedures and is incorporated herein in its entirety . herbicide resistance and resistance to another herbicide [0091 ] In the present invention , hybrids resistant to her group or insect resistance , such as resistance to white bicides such as, as but not limited to , ACC inhibiting sugarcane aphid as described herein was desirous, then herbicides , could be created by crossing parents in a pro US 2018 /0346920 A1 Dec. 6 , 2018 duction area where one or both parents contain the ACC hybrid , wherein at least one ancestor of the sorghum hybrid herbicide resistance trait . These parents , or parent, thereby comprises an ACC resistant gene from germplasm desig confer onto the resulting hybrid the herbicide resistance nated BTX430 - CHR - ACC1 or or BTX430 - CHR - ACC2 or through genetic combination of the parent lines to the F BTX430 -CHR -ACC4 . In some embodiments , the ACC hybrid seed . The present invention is not limited to the elite resistant herbicide gene comprises one or more of the parent sorghum lines listed , and one skilled in the art will mutations disclosed herein , for example the gene comprise recognize that any elite sorghum line would be equally at least one mutation described herein , or at least two amenable to the compositions and methods as described herein . The present invention is not limited to field produc mutations described herein or at least three mutations tion of hybrid seed , and one skilled in the art will recognize described herein . In addition , the ACC resistant herbicide that hybrid seed production could be conducted in any gene is at least 70 % homologous, at least 80 % homologous , environment where sorghum plants can be grown and pro at least 85 % homologous , at least 90 % homologous , at least duce seed . 95 % homologous , at least 97 % homologous , or at least 99 % [0092 ] In the present invention , open pollinated sorghum homologous to the ACC resistant herbicide gene as found in varieties (OPV ) could be created that are resistant to , but not the germplasm BTX430 -CHR - ACC1 or BTX430 - CHR limited to , ACC inhibitor herbicides . Open pollinated vari ACC2 or BTX430 -CHR - ACC4 . In some embodiments , the eties are different from inbred lines in that they are devel ACC resistant herbicide gene is at least 70 % homologous, at oped to be grown and the caryopses produced used as both least 80 % homologous, at least 85 % homologous, at least grain and seed for future plant propagation . Grain uses may 90 % homologous , at least 95 % homologous, at least 97 % include, but are not limited to , animal feed , human food , homologous , or at least 99 % homologous to the ACC ethanol production , and organic chemical production . Open resistantherbicide gene as found in the germplasm BTX430 pollinated variety breeding programs combine desirable CHR - ACC1, BTX430 - CHR - ACC2 or BTX430 - CHR traits; in the present invention , the desirable trait being plant ACC4 , such as germplasm comprising a tryptophan to resistance to ACC herbicides, by crossing a plant that cysteine at amino acid position aligning with Trp 1999 of the contains the trait with an OPV , followed by selection of CT Domain of the ACC gene ; or a tryptophan to serine plants with the desired trait . amino acid substitution at an amino acid position aligning [0093 ] In one embodiment, the present invention provides with Trp1999 of the CT domain of the ACC gene; or an a sorghum germplasm that confers resistance to inhibition by alanine to valine at amino acid position aligning with ACC herbicides, singly or in conjunction with other pest Ala2004 of the CT Domain of the ACC gene ; or a tryptophan resistance traits , for example insect tolerance to white sug to serine at amino acid position aligning with Trp2027 of the arcane aphid (Melanaphis sacchari (Zehntner ) (J . S . Arm CT Domain of the ACC gene. strong et al. , J . of Econ Entomol. Vol 108 , Pages 576 -582 , [0095 ] In another embodiment, the present invention pro incorporated herein in its entirety ) . In some embodiments , vides ACC herbicide resistance in sorghum plants compris for example , a sorghum hybrid whose germplasm comprises ing a mutation in the ACC gene found in the germplasm a synthetic cryl Ac gene from Bacillus thuringiensis ( Bt) is from BTX430 - CHR -ACC1 or BTX430 - CHR - ACC2 or introgressed into a sorghum line whose germplasm confers BTX430 -CHR - ACC4, wherein the mutation is incorporated resistance to ACC herbicides. In some embodiments , for inteinto elite sorghum varieties through gene editing technology, example , a sorghum hybrid whose germplasm comprises a thereby providing for the development of herbicide tolerant resistance to the fungal leaf disease Anthracnose (Colletotri sorghum crop hybrids that will tolerate the use of ACC chum graminicola ) is introgressed into a sorghum line inhibiting herbicides for weed control. In some embodi whose germplasm confers resistance to ACC herbicides . As ments, the ACC resistant herbicide gene is at least 70 % well , the incorporation of ACC herbicide resistance and homologous, at least 80 % homologous , at least 85 % insect resistance is accomplished via plant transgenesis into homologous , at least 90 % homologous, at least 95 % the same sorghum hybrid . One skilled in the art will recog homologous, at least 97 % homologous , or at least 99 % nize the various techniques as described herein that are homologous to the ACC resistant herbicide gene as found in applicable to the incorporation of two or more resistance the germplasm BTX430 -CHR - ACC1 or BTX430 - CHR attributes into the same sorghum plant. ACC2 or BTX430 -CHR - ACC4. In some embodiments, the [ 0094 ] In one embodiment, the present invention provides ACC resistant herbicide gene comprises one or more of the ACC herbicide resistance in sorghum plants comprising a mutations disclosed herein , for example the gene comprise mutation in the ACC gene found in the germplasm from at least one mutation described herein , or at least two BTX430 - CHR - ACC1 or BTX430 - CHR - ACC2 or BTX430 mutations described herein or at least three mutations CHR - ACC4 , wherien the mutation is incorporated into elite described herein . In addition , the ACC resistant herbicide sorghum varieties through plant breeding and selection , gene is at least 70 % homologous, at least 80 % homologous , thereby providing for the development of herbicide tolerant at least 85 % homologous , at least 90 % homologous , at least sorghum crop hybrids that will tolerate the use of ACC 95 % homologous , at least 97 % homologous, or at least 99 % inhibiting herbicides for weed control. Deployment of this homologous to the ACC resistant herbicide gene as found in herbicide tolerance trait in the aforementioned hybrids the germplasm BTX430 -CHR - ACC1 or BTX430 - CHR allows use of these herbicides to controlmonocot weeds that ACC2 or BTX430 - CHR - ACC4 , such as germplasm com grow in the presence of these crops. In some embodiments, prising a tryptophan to cysteine at amino acid position the incorporation of the ACC resistance germplasm into elite aligning with Trp1999 of the CT Domain of the ACC gene ; lines is via introgression , or classical breeding methods . In or a tryptophan to serine amino acid substitution at an amino some embodiments , the incorporation of the ACC resistance acid position aligning with Trp1999 of the CT domain of the gene into elite lines is via heterologous gene transgenesis . In ACC gene ; or an alanine to valine at amino acid position some embodiments , the invention provides a sorghum aligning with Ala2004 of the CT Domain of the ACC gene ; US 2018 /0346920 A1 Dec . 6 , 2018 11 or a tryptophan to serine at amino acid position aligning with equally preferred . Any of the methods disclosed herein are Trp2027 of the CT Domain of the ACC gene . contemplated to generate the ACC inhibitor herbicide tol [ 0096 ] In some embodiments , ACC herbicide resistant erant sorghum plant or plant part of the invention . germplasm is introgressed into an elite sorghum line using [0102 ] In one embodiment, the present invention provides classic breeding techniques. Examples of classical breeding use of a transgene comprising a heterologous gene such as methods for sorghum can be found in , for example , Sleper a gene encoding a mutant ACC protein for providing the and Poehlman , 2006 , Breeding Field Crops, Fifth Edition , selected agronomic trait of ACC herbicide resistance . In one Blackwell Publishing , incorporated herein in its entirety . embodiment, the transgene comprises a mutant ACC gene as 10097 ] In one embodiment, the ACC herbicide resistant found in the germplasm designated BTX430 - CHR - ACC1 or germplasm is introgressed into a sorghum plant that provides BTX430 - CHR - ACC2 or BTX430 - CHR -ACC4 . In some food for human consumption . In some embodiments , the embodiments , the transgene comprises one or more of the ACC herbicide resistant germplasm is introgressed into mutations disclosed herein . In addition , the transgene is at sorghum plants that provide food for livestock ( e . g . , poultry , least 70 % homologous , at least 80 % homologous , at least cattle , swine, sheep , etc ) . In some embodiments, the ACC 85 % homologous, at least 90 % homologous, at least 95 % herbicide resistant germplasm is introgressed into sorghum homologous, at least 97 % homologous, or at least 99 % plants that are used in industrial processes such as ethanol homologous or is 100 % identical to the ACC resistant production , the production of organic chemicals or energy herbicide gene as found in the germplasm BTX430 - CHR production from direct combustion of sorghum plant mate ACC1, or BTX430 - CHR - ACC2 or BTX430 -CHR -ACC4 rials . In one embodiment, the ACC herbicide resistant gene ( e . g . , the nucleotide sequences set out as any one of SEQ ID is introduced into the plant genome via transgenesis using NOS : 1 - 5 ) . In some embodiments, the ACC resistant herbi vectors and technologies known in the art or by classical cide gene is at least 70 % homologous , at least 80 % homolo breeding gous, at least 85 % homologous, at least 90 % homologous , at 10098 ] In some embodiments , the present invention pro least 95 % homologous, at least 97 % homologous , or at least vides an ACC resistant germplasm of a sorghum plant part 99 % homologous or is 100 % identical to the ACC resistant of germplasm BTX430 -CHR - ACC1 or BTX430 -CHR herbicide gene as found in the germplasm BTX430 -CHR ACC2 or BTX430 - CHR -ACC4 , and said sorghum plant part ACC1 or BTX430 -CHR - ACC2 or BTX430 -CHR - ACC4 , is one or more of a pollen , an ovule , a tissue , a pod , a seed , such as germplasm comprising a tryptophan to cysteine at and a cell . In one embodiment , the present invention pro amino acid position aligning with Trp 1999 of the CT Domain vides an F? hybrid whose germplasm comprises an ACC of the ACC gene; or a tryptophan to serine amino acid resistance gene as described herein . In some embodiments , substitution at an amino acid position aligning with Trp 999 the F , hybrid is a cross between two elite sorghum lines , at of the CT domain of the ACC gene (SEQ ID NO : 7 or 8 ; least one of which contains a germplasm comprising an respectively ) , or an alanine to valine at amino acid position ACC resistance gene as described herein . aligning with Ala2004 of the CT Domain of the ACC gene 10099 ] The present invention is not limited to sorghum (SEQ ID NO : 9 ) ; or a tryptophan to serine at amino acid plants mutated with EMS . Within the scope of the present position aligning with Trp2007 of the CT Domain of the ACC invention are sorghum plants obtained by other mutation gene (SEQ ID NO : 10 ). methods, for example methods such as radiation and chemi [0103 ] Heterologous genes intended for expression in cal mutagens. Herbicide - resistant mutant plants can also be plants are first assembled in expression vectors containing a obtained by means of a process of selective pressure on cells heterologous gene and appropriate transcriptional and trans cultured with a herbicide and selection of resistant cells to lational control elements , methods of which are well known generate a herbicide -resistant plant. Details of mutation and to those skilled in the art . Methods include in vitro recom breeding methods can be found in “ Principles of Cultivar binant DNA techniques , synthetic techniques, and in vivo Development” Fehr , 1993 , Macmillan Publishing Company, genetic recombination . Exemplary techniques are widely the disclosure of which is included herein by reference . described in the art ( See e . g . , Sambrook . et al . ( 1989 ) Someone skilled in the art may also be able to create Molecular Cloning , A Laboratory Manual, Cold Spring mutations by exposing seeds or callus to , but not limited to , Harbor Press , Plainview , N . Y . , and Ausubel, F . M . et al. the following: ion beams, cosmic radiation , X - ray radiation ( 1989 ) Current Protocols in Molecular Biology , John Wiley and gamma radiation . & Sons, New York , N . Y ., herein incorporated by reference ). f0100 ) Gene editing methods can also be used to develop [0104 ] In general, these vectors comprise a nucleic acid mutations that creating traits with herbicide tolerant func sequence encoding a heterologous gene operably linked to a tions . Sequence - specific nuclease can be designed to target promoter and / or other regulatory sequences ( e . g . , enhancers , sorghum ACCase carboxyltransferase domain (CT domain ). polyadenylation signals , etc .) required for expression in a Once the nuclease make the double strand break on the plant. target sequence in a living cell, the cell' s DNA repair 0105 ] Promoters include , but are not limited to , consti machinery will find the break and try to fix it. Mutations are tutive promoters , tissue - , organ - , and developmentally spe created when the repair is not perfect . It could be a deletion cific promoters , and inducible promoters . Examples of pro or insertion by DNA repair through non -homology end moters include , but are not limited to ; constitutive promoter joining (NHEJ ) pathway . Or specific sequence modification 35S of cauliflower mosaic virus; a wound - inducible pro (alteration can be obtained when DNA repaired through moter from tomato , leucine amino peptidase (Chao et al . , homologous recombination (HR ) pathway with designed 1999 , Plant Physiol 120 :979 - 992 , herein incorporated by sequence (donor ) applied . reference ) ; a chemically -inducible promoter from tobacco , [0101 ] The approaches to modify the nucleotide sequence Pathogenesis -Related 1 ( induced by salicylic acid and ben of the sorghum ACC gene to confer ACC inhibiting herbi zothiadiazole- 7 - carbothioic acid S -methyl ester ) ; a heat cides tolerance disclosed herein are equally relevant and shock promoter (U . S . Pat. No . 5 ,187 ,267 , herein incorpo US 2018 /0346920 A1 Dec . 6 , 2018 rated by reference ) ; a tetracycline - inducible promoter ( U . S . phosphinothricin (White et al ., 1990 . Nucl Acids Res . Pat . No . 5 ,057 , 422 , herein incorporated by reference ); and 18 : 1062 ) , the hph gene which confers resistance to the seed - specific promoters . antibiotic hygromycin (Blochlinger and Diggelmann , 1984 , [0106 ] The expression cassettes may further comprise any Mol. Cell. Biol . 4 :2929 , incorporated herein by reference) , sequences required for expression of mRNA . Such and the dhfr gene that confers resistance to methotrexate sequences include , but are not limited to transcription ter (Bourouis et al. , 1983 , EMBO J. , 2 : 1099 , incorporated minators , enhancers such as introns , viral sequences , and herein by reference ). sequences intended for the targeting of the gene product to [0112 ] In some embodiments , the Ti (T -DNA ) plasmid specific organelles and cell compartments . vector is adapted for use in an Agrobacterium mediated [0107 ] A variety of transcriptional terminators are avail transfection process such as in U . S . Pat. No . 6 ,369 , 298 able for use in expression of sequences using the promoters ( sorghum ) , and U . S . Pat. Nos. 5 , 981 , 839 , 6 , 051, 757 , 5 , 981, such as those disclosed herein . Transcriptional terminators 840 , 5 ,824 , 877 and 4 , 940 ,838 all of which are incorporated are responsible for the termination of transcription beyond by reference herein in their entireties . Construction of the transcript and its correct polyadenylation . Appropriate recombinant Ti and Ri plasmids in general follows methods transcriptional terminators and those which are known to typically used with more common vectors, such as pBR322 . function in plants include , but are not limited to , the CaMV Additional use can be made of accessory genetic elements 35S terminator, the tml terminator , the pea rbcS E9 termi sometimes found with the native plasmids and sometimes nator , and the nopaline and octopine synthase terminator constructed from foreign sequences . These may include , but ( Odell et al. , 1985 , Nature 313 : 810 ; Rosenberg et al. , 1987 , are not limited to , structural genes for antibiotic resistance as Gene , 56 : 125 ; Guerineau et al. , 1991, Mol. Gen . Genet . selection genes . 262: 141; Proudfoot, 1991, Cell , 64 :671 ; Sanfacon et al. , [0113 ]. There are two systems of recombinant Ti and Ri 1991 , Genes Dev . 5 : 141 all of which are incorporated herein plasmid vector systems now in use . The first system is called by reference ) . the " cointegrate ” system . In this system , the shuttle vector 0108 ] In some embodiments , constructs for expression of containing the gene of interest is inserted by genetic recom the heterologous gene of interest include one or more of bination into a non -oncogenic Ti plasmid that contains both sequences found to enhance gene expression from within the the cis -acting and trans -acting elements required for plant transcriptional unit . These sequences can be used in con transformation as , for example , in the PMLJ1 shuttle vector junction with the nucleic acid sequence of interest to and the non -oncogenic Ti plasmid pGV3850 . The use of increase expression in plants . Various intron sequences have T -DNA as a flanking region in a construct for integration been shown to enhance expression , particularly in mono into a Ti - or Ri- plasmid has been described in EPO No . cotyledonous cells . Intron sequences have been routinely 116 ,718 and PCT Application Nos. WO 84 /02913 , 02919 incorporated into plant transformation vectors , typically and 02920 ; Herrera - Estrella , 1983 , Nature 303: 209 - 213 ; within the non - translated leader. Fraley et al. , 1983 , Proc. Natl. Acad . Sci, USA 80 :4803 [0109 ] In some embodiments , a construct for expression 4807 ; all of which are herein incorporated by reference . of the heterologous nucleic acid sequence of interest also [0114 ]. The second system is called the “ binary ” system in includes a regulator such as a nuclear localization signal which two plasmids are used and the gene of interest is (Kalderon et al. , 1984 , Cell 39 :499 ; a plant translational inserted into a shuttle vector containing the cis -acting ele consensus sequence ( Joshi, 1987 , Nucleic Acids Research ments required for plant transformation . The other necessary 15 :6643 ) , an intron ( Luehrsen and Walbot, 1991 , Mol. Gen . functions are provided in trans by the non -oncogenic Ti Genet . 225 : 81) , and the like, operably linked to the nucleic plasmid as exemplified by the pBIN19 shuttle vector and the acid sequence encoding an heterologous gene . non -oncogenic Ti plasmid PAL4404 . Some of these vectors [0110 ] In preparing the construct comprising the nucleic are commercially available . acid sequence encoding an heterologous gene , or encoding [0115 ] In some embodiments, the nucleic acid sequence of a sequence designed to decrease heterologous gene expres interest is targeted to a particular locus on the plant genome . sion , various DNA fragments can be manipulated so as to Site - directed integration of the nucleic acid sequence of provide for the DNA sequences in the desired orientation interest into the plant cell genome may be achieved by , for (e .g ., sense or antisense ) and , as appropriate , in the desired example , homologous recombination using Agrobacterium reading frame. For example, adapters or linkers can be derived sequences. Generally , plant cells are incubated with employed to join the DNA fragments , or other manipula a strain of Agrobacterium which contains a targeting vector tions can be used to provide for convenient restriction sites , in which sequences that are homologous to a DNA sequence removal of superfluous DNA , removal of restriction sites, inside the target locus are flanked by Agrobacterium trans and the like . For this purpose , in vitro mutagenesis , primer fer -DNA ( T -DNA ) sequences, as previously described ( U . S . repair , restriction , annealing , resection , ligation , and the like Pat . No. 5 ,501 , 967 herein incorporated by reference ). Oneof is preferably employed , where insertions, deletions or sub skill in the art knows that homologous recombination may stitutions ( e . g . , transitions and transversions ) are involved . be achieved using targeting vectors that contain sequences [ 0111 ] Numerous transformation vectors are available for that are homologous to any part of the targeted plant gene , plant transformation . The selection of a vector for use will whether belonging to the regulatory elements of the gene or depend upon the preferred transformation technique and the the coding regions of the gene . Homologous recombination target species for transformation . For certain target species, may be achieved at any region of a plant gene so long as the different antibiotic or herbicide selection markers are pre nucleic acid sequence of regions flanking the site to be ferred . Selection markers used routinely in transformation targeted is known . Agrobacterium tumefaciens is a common include the nptll gene which confers resistance to kanamycin soil bacterium that causes crown gall disease by transferring and related antibiotics (Messing and Vierra , 1982 , Gene 19 : some of its DNA to the plant host. The transferred DNA 259 , the bar gene which confers resistance to the herbicide ( T -DNA ) is stably integrated into the plant genome, where US 2018 /0346920 A1 Dec. 6 , 2018 13 its expression leads to the synthesis of plant hormones and 5 ,451 , 513 and 5 , 545, 817 ) all of which are incorporated thus to the tumorous growth of the cells . A putative macro herein by reference in their entireties ). molecular complex forms in the process of T -DNA transfer [0120 ] The basic technique for chloroplast transformation out of the bacterial cell into the plant cell. involves introducing regions of cloned plastid DNA flanking [0116 ] In some embodiments , the nucleic acids as dis a selectable marker together with the nucleic acid encoding closed herein are utilized to construct vectors derived from the sequences of interest into a suitable target tissue ( e . g . , plant ( + ) RNA viruses ( e . g ., brome mosaic virus , tobacco using biolistics or protoplast transformation with calcium mosaic virus, alfalfa mosaic virus , cucumber mosaic virus , chloride or PEG ) . The 1 to 1 . 5 kb flanking regions, termed tomato mosaic virus , and combinations and hybrids thereof) . targeting sequences, facilitate homologous recombination Generally , the inserted heterologous polynucleotide can be with the plastid genome and thus allow the replacement or expressed from these vectors as a fusion protein ( e . g . , coat modification of specific regions of the plastome. Initially , protein fusion protein ) or from its own subgenomic pro point mutations in the chloroplast 16S rRNA and rps12 moter or another promoter. Methods for the construction and genes conferring resistance to spectinomycin and / or strep use of such viruses are described in U . S . Pat. Nos . 5 ,846 , tomycin are utilized as selectablemarkers for transformation 795 ; 5 ,500 , 360 ; 5 , 173 ,410 ; and 5 , 965 ,794 ; all of which are (Svab et al. , 1990 , Proc. Natl . Acad . Sci. , 87 :8526 ) ; Staub incorporated herein by reference . and Maliga , 1992 , Plant Cell , 4 : 39 , all of which are incor [0117 ] In some embodiments , a heterologous nucleic acid porated herein by reference ) . The presence of cloning sites sequence of interest comprising a mutant ACC transgene , between these markers allows creation of a plastid targeting for example, as found in the germplasm designated vector introduction of foreign DNA molecules ( Staub and BTX430 -CHR - ACC1 or BTX430 -CHR - ACC2 or BTX430 Maliga , 1993 , EMBO J ., 12 :601 ) . Substantial increases in CHR -ACC4 , is introduced directly into a plant. In some transformation frequency are obtained by replacement of the embodiments , the transgene is at least 70 % homologous , at recessive rRNA or r -protein antibiotic resistance genes with least 80 % homologous , at least 85 % homologous, at least a dominant selectable marker, the bacterial aadA gene 90 % homologous, at least 95 % homologous , at least 97 % encoding the spectinomycin -detoxifying enzyme aminogly homologous , or at least 99 % homologous or 100 % identical coside- 3 - adenyltransferase (Svab and Maliga , 1993 , Proc . to the ACC resistant herbicide gene as found in the germ Natl . Acad . Sci . , 90 : 913 ) . Other selectable markers useful plasm BTX430 -CHR - ACC1 or BTX430 -CHR -ACC2 or for plastid transformation are known in the art and encom BTX430 - CHR - ACC4 (e . g ., any one of SEQ ID NOS : 2 - 5 ). passed within the scope of the present invention . Plants In some embodiments , the transgene is at least 70 % homolo homoplasmic for plastid genomes containing the two nucleic gous , at least 80 % homologous , at least 85 % homologous , at acid sequences separated by a promoter of the present least 90 % homologous , at least 95 % homologous , at least invention are obtained , and are preferentially capable of high 97 % homologous , or at least 99 % homologous or 100 % expression of RNAs encoded by the DNA molecule . identical to the ACC resistant herbicide gene as found in the [0121 ] In one embodiment, vectors useful in the practice germplasm BTX430 - CHR - ACC1 or BTX430 - CHR - ACC2 of the present invention are microinjected directly into plant or BTX430 - CHR - ACC4 , such as germplasm comprising a cells ( Crossway , 1985 , Mol. Gen . Genet , 202 : 179 ) . In some tryptophan to cysteine at amino acid position aligning with embodiments , the vector is transferred into the plant cell by Trp 1999 of the CT Domain of the ACC gene ; or a tryptophan using polyethylene glycol (Krens et al. , 1982 , Nature , 296 : to serine amino acid substitution at an amino acid position 72 ) fusion of protoplasts with other entities such as mini aligning with Trp1999 of the CT domain of the ACC gene cells , cells, lysosomes or other fusible lipid - surfaced bodies (SEQ ID NO : 7 or 8 ; respectively ) , or an alanine to valine ( Fraley et al. , 1982 , Proc . Natl. Acad . Sci ., USA , 79 : 1859 ) ; at amino acid position aligning with Ala2004 of the CT and protoplast transformation (EP O 292 435 ) ; direct gene Domain of the ACC gene ( SEQ ID NO : 9 ); or a tryptophan transfer ( Paszkowski et al. , 1984 , EMBO J . , 3 : 2717 . In some to serine at amino acid position aligning with Trp2027 of the embodiments , the vector may also be introduced into the CT Domain of the ACC gene (SEQ ID NO : 10 ) . plant cells by electroporation . ( Fromm , et al . , 1985 , Proc . [0118 ] One vector useful for direct gene transfer tech Natl. Acad . Sci. USA 82 : 5824 ; Riggs and Bates ., 1986 , Proc. niques in combination with selection by the herbicide Basta Natl . Acad . Sci . USA 83 :5602 ) . In this technique, plant ( or phosphinothricin ) is a modified version of the plasmid protoplasts are electroporated in the presence of plasmids PCIB246 , with a CaMV 35S promoter in operational fusion containing the gene construct . Electrical impulses of high to the E . coli GUS gene and the CaMV 35S transcriptional field strength reversibly permeabilize biomembranes allow terminator (WO 93/ 07278 , herein incorporated by refer ing the introduction of the plasmids. Electroporated plant ence ). protoplasts reform the cell wall, divide , and form plant [0119 ] Once a nucleic acid sequence encoding the heter callus . ologous gene is operatively linked to an appropriate pro [0122 ] In addition to direct transformation , in some moter and inserted into a suitable vector for the particular embodiments , the vectors comprising a nucleic acid transformation technique utilized ( e . g . , one of the vectors sequence encoding a heterologous gene are transferred using described above ) , the recombinant DNA described above Agrobacterium -mediated transformation (Hinchee et al. , can be introduced into the plant cell in a number of art 1988 , Nature Biotechnology , 6 : 915 ; Ishida et al. , 1996 , recognized ways . Those skilled in the art will appreciate that Nature Biotechnology 14 :745 , all of which are herein incor the choice of method depends on the type of plant targeted porated by reference ) . Agrobacterium is a representative for transformation . In some embodiments , the vector is genus of the gram - negative family Rhizobiaceae . Its species maintained episomally . In some embodiments , the vector is are responsible for plant tumors such as crown gall and hairy integrated into the genome. In some embodiments, direct root disease . In the dedifferentiated tissue characteristic of transformation in the plastid genome is used to introduce the the tumors , amino acid derivatives known as opines are vector into the plant cell ( for example , see U . S . Pat. Nos. produced and catabolized . The bacterial genes responsible US 2018 /0346920 A1 Dec . 6 , 2018 14 . for expression of opines are a convenient source of control ACCase mutations developed through gene editing can be elements for chimeric expression cassettes . Heterologous but not limited to W1999C , W1999S , A2004V , W2027S . genetic sequences ( e . g . , nucleic acid sequences operatively linked to a promoter of the present invention ) can be [0129 ] Tissue culture, transformation and regeneration of introduced into appropriate plant cells , by means of the Ti sorghum have been reported by several groups since it was plasmid of Agrobacterium tumefaciens (previously first transformed in 1991 . Although sorghum has been described ) . The Ti plasmid is transmitted to plant cells on successfully transformed by both Agrobacterium and biolis infection by Agrobacterium tumefaciens , and is stably inte tic methods , the efficiency of transformation is very low grated into the plant genome (Schell , 1987 , Science , 237 : (< 10 % ) causing sorghum to be classified as a recalcitrant 1176 ) . Species that are susceptible to infection by Agrobac crop for both tissue culture and genetic transformation . terium may be transformed in vitro . Transformation methods Increasing the transformation efficiency to higher levels ( as for producing transgenic sorghum plants using Agrobacte observed in other monocot crop plants such as corn , rice and rium -mediated transformation are provided in U . S . Pat. No . sugarcane ) would be valuable to more rapidly genetically 6 , 369, 298 . improve the crop . An improved transformation system [0123 ] In some embodiments , the vector is introduced would reduce time, cost, and resources necessary to evaluate through ballistic particle acceleration (U . S . Pat . No . 4 ,945 , genetic elements in sorghum and to generate transgenic 050 ; Casas et al. , 1993 , Proc . Natl . Acad . Sci. USA plants . Many variables can influence sorghum transforma 90 : 11212 , all references are incorporated herein in their tion efficiency . Use of an efficient selection scheme to entireties ) . identify events is a critical variable for success . Several [0124 ] In some embodiments , after selecting for trans selectable maker genes, including Nptil , hptII , bar and pmi formed plant material that can express a heterologous gene have previously been tested in sorghum using a range of encoding a heterologous protein or variant thereof, whole plants are regenerated . Plant regeneration from cultured promoters (i . e . CaMV35S , riceActin1, maizeUbil , maize protoplasts is described in Evans et al. , Handbook of Plant ADH , etc . ) Cell Cultures, Vol. 1 : (MacMillan Publishing Co . New York , [0130 ] In some embodiemnts , sorghum transformation ( 1983 ) ; Vasil I . R . (ed .) , Cell Culture and Somatic Cell efficiency could be improved by replacing the yeast based Genetics of Plants , Acad . Press , Orlando , Vol. I , ( 1984 ) and promoter with a stronger plant promoter. In general , For Vol . III , (1986 ), incorporated herein by reference in their transforming sorghum , callus materials were produced from entireties. It is known that many plants can be regenerated immature sorghum as described by Gurel et al. , 2009 . Calli from cultured cells or tissues including , but not limited to , all are transferred to fresh media every three weeks until the major species of sugarcane , sugar beet , cotton , fruit and entire callus has turned into compact and friable callus . other trees , legumes and vegetables , and monocots ( e . g ., the Three week old calli are subcultured into 3 - 4 mm diameter plants described above ) . Means for regeneration vary from pieces and placed on medium that contains compounds to species to species of plants , but generally a suspension of maintain a prescribed osmotic level for 4 h prior bombard transformed protoplasts containing copies of the heterolo ment. Approximately 30 calli were placed at the center of a gous gene is first provided . Callus tissue is formed and Petri dish ( 15x90 mm ) containing osmotic medium and shoots may be induced from callus and subsequently rooted . stored for 4 - 5 hours under a prescribed light treatment prior [ 0125 ] Alternatively , embryo formation can be induced to bombardment. Plasmid DNA , can be coated onto 0 . 6 um from the protoplast suspension . These embryos germinate diameter gold particles as described by Carlson et al. , 2007 and form mature plants . The culture media will generally and delivered into calli using a gas powered biolistic deliv contain various amino acids and hormones, such as auxin ery system . Bombardmentmay be carried out with a system and cytokinins shoots and roots normally develop simulta configuration as described by Carlson , et al, 2007 . Follow neously. Efficient regeneration will depend on the medium , ing bombardment, the cultures are transferred onto medium on the genotype , and on the history of the culture . The and incubated under 28° C . for 4 - 6 days and then were reproducibility of regeneration depends on the control of transferred to medium containing a selective agent and these variables . cultured under long -days with fluorescent lighting for 14 [0126 ] In some embodiments , after selecting for trans days at 28° C . The surviving calli can selected with two formed plant material that can express a heterologous gene higher concentration of a selective agent at 14 days interval. encoding a heterologous protein or variant thereof, whole All the transgenic calli events can regenerated and devel plants are regenerated . Transformation efficiency is mea oped into a whole plant. sured as the percentage of regenerated resistant events out of [0131 ] In some embodiments , a method that employs a the total number of plant material used for transformation . strong plant promoter in combination with other promoters , 10127 ] In some embodiments , transgenic plants were in a preferred embodiment, this other promoter would be regenerated from green callus after ballistic particle accel Nptll , could significantly outperform a yeast promoter and eration ( U . S . Pat. No. 6 , 486 ,384 ) . High frequency sorghum NptIl combination . In some embodiments , the strong plant transformation procedure is developed . In some embodi promoter transformation efficiency ( TE ) is expected to ments, the transformation efficiency is at least 25 % , at least exceed 25 % with TE levels potentially as high as 55 % . This 30 % , at least 35 % , at least 40 % , at least 45 % , at least 50 % , frequency puts sorghum transformation on par with other or greater than 50 % . In some embodiments , the regenerated important monocot crops. In addition to being more efficient plants are resistant to ACCase inhibitor herbicide. in producing numbers of events , those events produced with [0128 ] Herbicide tolerant mutations can be developed strong plant promoter and NptII combination were also able through gene editing technology . In some embodiments , to be identified approximately 2 weeks earlier than events sequence -specific nucleases used for gene editing can be produced with yeast promoter and Nptil . The size (biomass ) ZFN , TALEN , CRISPR /Cas9 . In some embodiments , of the selected callus averaged 5 - fold larger with stronger US 2018 /0346920 A1 Dec. 6 , 2018 promoter and NptII than with yeast promoter and NptII , (0139 ] In one embodiment, the present invention provides allowing the earlier identification and for scientists to regen methods for controlling weeds in a field of any ACC erate more plants per event . herbicide resistant sorghum plants include hybrid sorghum [ 0132 ] Segregating plant populations must be screened to crop plants . In some embodiments , controlling the weeds identify progeny that contain desirable traits , resistance to comprises applying an ACC herbicide to said field of ACC inhibiting herbicides in the present invention . Plants sorghum plants, such that weed growth is inhibited but with desired traits can be screened via phenotypic and sorghum growth is not adversely affected . In some embodi genotypic methods . In the present invention , phenotypic ments , the ACC herbicide being applied is from the ary screening can be accomplished through the application of loxyphenoxypropionate (FOP ) , cyclohexanedione (DIM ) ACC inhibiting herbicides whereas genotypic screening can and phenylpyrazolin (DENS ) herbicide families including , be accomplished via genetic markers. but not limited to , clodinafop -propargyl (CAS RN 105512 [ 0133 ] In the present invention , marker identification of 06 - 9 ) : cyhalofop - butyl (CAS RN 122008 -85 - 9 ) ; diclofop plants containing the desired trait could be accomplished methyl (CAS RN 51338 -27 - 3 ); fenoxaprop - p - ethyl (CAS through the use of marker which include , but not limited to , RN 71283 - 80 - 2 ) ; fluazifop - P - butyl (CAS RN 79241 - 46 - 6 ) ; simple sequence repeat (SSR ) , single nucleotide polymor quizalofop - p - ethyl (CAS RN 100646 - 51- 3 ) ; quizalofop - p phism (SNP ) , genetic insertions and / or deletions. One (CAS RN 94051- 08 - 8 ) ; haloxyfop (CAS RN 69806 - 34 - 4 ) ; embodiment of the present invention might employ Kom haloxyfop - ethoxyethyl (CAS RN 87237 - 48 - 7 ) ; haloxyfop petitive Allele Specific PCR (KASP ) DNA markers devel etotyl (CAS RN 87237 -48 - 7 ) ; haloxyfop - R -methyl (CAS oped for all SNPs associated with the desired mutation . RN 72619 - 32 - 0 ) ; metamifop (CAS RN 256412 - 89 - 2 ) ; [ 0134 ] One skilled in the art could employ KASP by a real propaquizafop (CAS RN 111479 - 05 - 1 ) ; alloxydim (CAS time PCR allelic discrimination assay using a thermocycler RN 55634 - 91- 8 ) ; butroxydim (CAS RN 138164 - 12 - 2 ) ; and amplified PCR gels . The result of the allelic discrimi cycloxydim (CAS RN 101205 - 02 - 1 ); clethodim (CAS RN nation could be determined by endpoint detection using 99129 -21 - 2 ) ; profoxydim (CAS RN 139001 -49 - 3 ) ; sethoxy methods such as , but not limited to , fluorescence detection . dim (CAS RN 74051 - 80 - 2 ) ; tepraloxydim (CAS RN Comparison of KASP assay results between DNA extracted 149979 - 41 - 9 ) ; tralkoxydim (CAS RN 87820 - 88 - 0 ) ; pinoxa from plants that might contain the mutation and a wild type den (CAS RN 243973 -20 - 8 ). In some embodiments , the allow for the confirmation of presence or absence of the ACC herbicide being applied comprises a combination of desired mutation . This approach can be used on calli and compounds from both FOP and DIM ACC herbicide fami plants . lies as disclosed herein . However, the present application is [0135 ] Phenotypic screening could be accomplished not limited to the ACC herbicide used , and a skilled artisan through the application of herbicides from the family of the will appreciate that new ACC herbicides are being discov desired resistance trait ; in the present invention resistance to ered at any given time that inhibit the ACC enzyme. ACC inhibiting herbicides is the desired trait . In the present [0140 ] As such , one embodiment of the present invention invention , if plants from a segregating population created by provides a sorghum germplasm that contains altered ACC one of the methods described above are exposed to herbi genes and proteins . In some embodiments , the present cides from the FOP, DIM , or DEN herbicide groups, plants invention provides for the use ofACC herbicides in fields of containing the desired mutation are expected to survive . hybrid sorghum crop plants to reduce the amount of [ 0136 ] One skilled in the art will recognize that the unwanted vegetation present in said crop field , wherein said appropriate application dose is important for developing hybrid sorghum germplasm comprises an altered ACC plants with the appropriate level of resistance . In the present enzyme that confers resistance to ACC herbicides and said invention , plants that contain the mutations that result in weed plants are ACC herbicide susceptible . plants that are resistant to ACC inhibiting herbicides could [0141 ] As such , one embodiment of the present invention be detected by applying the recommend rate of a herbicide provides a sorghum germplasm that contains altered ACC from the FOP, DIM , or DEN family based on the herbicide genes and proteins . In some embodiments , the present manufacturer ' s recommendation for controlling shattercane invention provides for the use of ACC herbicides in fields of ( Sorghum bicolor ). In the present invention , the herbicide hybrid sorghum crop plants to reduce the amount of mono application rate might also be increased to twice the rate or cot weed plants present in said crop field , wherein said might also be increased to four times the rate as recom hybrid sorghum germplasm comprises an altered ACC mended by the herbicide manufacturer for control of shat enzyme that confers resistance to ACC herbicides and said tercane (Sorghum bicolor ) . weed plants are ACC herbicide susceptible . [ 0137 ] One skilled in the art will recognize that the 10142 ] In one embodiment, the removal of unwanted herbicide application may occur at various stage of plant or vegetation from a sorghum growing area that comprises the cell development. In the present invention , the desired presence of one or more sorghum plants that are resistant to herbicide is from the FOP , DIM or DEN family , and could ACC inhibiting herbicides and applying one or more ACC be applied to the media that is used to produce callus or other inhibitor herbicide ( s ) alone or in combination with one or artificial plant propagation media . more herbicide ( s ) that do ( es ) not belong to the class of ACC [0138 ] Alternatively , the desired herbicide could be inhibitor herbicides (non - ACC inhibitor herbicides ), and applied to irrigation water or hydroponic solutions used to wherein the application of the herbicides as defined under propagate plants , or could be applied directly to the foliage takes place jointly or simultaneously , or takes place at of plants being grown in soil or other media in a field , different times and /or in a plurality of portions (sequential greenhouse , or plant growth chamber . These plants may application ) . These applications may include a pre - emer range in age from the presence of a single leaf collar to gence applications followed by post -emergence applications physiological maturity, which is identified by the presence of or early post- emergence applications followed by medium a black layer at the base of the mature caryopsis . or late post - emergence applications. However , the present US 2018 /0346920 A1 Dec. 6 , 2018 16 application is not limited to the ACC herbicide used or the which are difficult to control. Here , the substances can be application timing or methods and a skilled artisan will applied for example, by the pre -sowing method , the pre appreciate that new ACC herbicides are being discovered at emergence method , or the post- emergence method , for any given time that inhibit the ACC enzyme. example jointly or separately . Preference is given , for [0143 ] In one embodiment, the removal of unwanted example , to application by the post -emergence method , in vegetation , or weeds, could from be from areas where hybrid particular to emerged harmful plants or unwanted vegeta sorghum seed is being produced from one or more parent tion . lines ( A - line or R - line ) that is resistant to ACC herbicides . These areas could include greenhouse growing areas and [0147 ] Examples ofweed species on which the application vessels or production fields . However , the present applica according to present invention act efficiently are , from tion is not limited to the ACC herbicide used , and a skilled amongst the monocotyledonous weed species, Avena spp ., artisan will appreciate that new ACC herbicides are being Alopecurus spp ., Apera spp ., Brachiaria spp . , Bromus spp . , discovered at any given time that inhibit the ACC enzyme . Digitaria spp . , Lolium spp. , Echinochloa spp ., Panicum 10144 ] In one embodiment, the removal of unwanted spp ., Phalaris spp ., Poa spp ., Setaria spp . and also Cyperus vegetation , or weeds, could from be from areas where species from the annual group , and , among the perennial sorghum grain is being produced from one or more hybrids species , Agropyron , Cynodon , Imperata and Sorghum and that are resistant to ACC herbicides . These areas could also perennial Cyperus species. include greenhouse growing areas and vessels or end user grain production fields. However, the present application is [ 0148 ] Field crops have been classically bred through not limited to the ACC herbicide used , and a skilled artisan techniques that take advantage of the plants method ( s ) of will appreciate that new ACC herbicides are being discov pollination . A plant is considered “ self- pollinating ” if pollen ered at any given time that inhibit the ACC enzyme. from one flower can be transmitted to the same or another [ 0145 ] In one embodiment, the present invention provides flower, whereas plants are considered " cross - pollinated ” if for an ACC herbicide resistant sorghum plants or a sorghum the pollen has to come from a flower on a genetically hybrid ( e . g ., F1, F2, F3 , F4, etc . ) whose germplasm confers different plant in order for pollination to occur . resistance to ACC herbicides and resistance to one or more [0149 ] Plants that are self -pollinated and selected over additional herbicides from one or more different herbicide many generations become homozygous at most , if not all , of groups . For example , additional herbicide groups used to their gene loci , thereby producing a uniform population of inhibit weed growth , include , but are not limited to , inhibi tors of lipid synthesis (e . g. , aryloxyphenoxypropionates, true breeding progeny . A cross between two homozygous cyclohexanodeiones , benzofuranes, chloro -carbonic acids , plants from differing backgrounds or two different homozy phosphorodithioates , thiocarbamates ), inhibitors of photo gous lines will produce a uniform population of hybrid synthesis at photosystem II (e . g. , phenyl- carbamates, plants that will more than likely be heterozygous at a number pyridazinones , triazines , triazinones , triazolinones , uracils , of the gene loci. A cross of two plants that are each amides , ureas , benzothiadiazinones, nitriles, phenyl- pyri heterozygous at a number of gene loci will produce a dines ) , inhibitors of photosynthesis at photosystem I ( e . g . , generation ofhybrid plants that are genetically different and bipyridyliums) , inhibitors of protoporphyrinogen oxidase are not uniform . ( e . g . , diphenylethers , N - phenylphthalimides, oxadiazoles , oxyzolidinediones , phenylpyrazoles , pyrimidindiones , thia [0150 ] Host plant resistance is the most effective and diazoles ) , inhibitors of carotenoid biosynthesis ( e . g ., economical approach to minimized the economic losses in pyridazinones , pyridinecarboxamides, isoxazolidinones, tri crops to pests . Crop producers select cultivars , hybrids or azoles ) , inhibitors of 4 -hydroxyphenyl - pyruvate - dioxy open pollinated varieties, to plant in their fields based on genase ( e . g ., callistemones , isoxazoles , pyrazoles, trike host plant resistances to protect their crops some expected tones) , inhibitors of EPSP synthase ( e. g. , glycines ), pests in their region . In a preferred embodiment , sorghum inhibitors of glutamine synthetase ( e . g ., phosphinic acids) , farmers would select to plant sorghum seed that can produce inhibitors of dihydropteroate synthase ( e . g . , carbamates ), plants that are resistant to ACC inhibiting herbicides . The inhibitors ofmicrotubule assembly ( e . g . , benzamides , ben application of this trait, resistance to ACC inhibiting herbi zoic acids, dinitroanilines, phosphoroamidates, pyridines ), cides by sorghum farmers facilitates the application of ACC inhibitors of cell division (e .g ., acetamides, chloroacet inhibiting herbicides over their fields as a means of control amides , oxyacetamides ) , inhibitors of cell wall synthesis ling unwanted grass species vegetation in their fields. A ( e. g ., nitriles , triazolocarboxamides) and inhibitors of auxin preferred application method would be a broadcast spray, or transport ( e . g ., phthalamates , semicarbazones ). In some nonselective nor non - directed spray , over the top of the embodiments , the present invention provides F1 hybrids entire field that contains both sorghum plants with resistance from elite sorghum lines that comprises resistance to one or to ACC inhibitin herbicides and unwanted vegetation . more ACC herbicides alone , or in conjunction with , herbi cide resistance to one or more of the aforementioned her [0151 ] All publications and patents cited in this disclosure bicide groups . However, the present application is not lim are incorporated by reference in their entirety . To the extent ited to the these non - ACC herbicides used , and a skilled the material incorporated by reference contradicts or is artisan will appreciate that new non - ACC herbicides are inconsistent with this specification , the specification will being discovered at any given time that could be combine supersede any such material. with herbicides that inhibit the ACC enzyme. [0152 ] The following examples are provided in order to [0146 ] The application of ACC inhibitor herbicides also demonstrate and further illustrate certain preferred embodi act efficiently on perennial weeds which produce shoots m ents and aspects of the present invention and are not to be from rhizomes, root stocks, and other perennial organs construed as limiting the scope thereof . US 2018 /0346920 A1 Dec . 6 , 2018 17

EXAMPLES these resistant calli and established as whole plants in the greenhouse . The plants regenerated from resistant calli (FP Example 1 : Plant Material, Isolation of Immature 8 , FP - 11 and FP - 12 ) through chemical mutagenesis of Embryos (IES ) , Production of Sorghum Callus and sorghum calli, showed dwarf plant phenotype and male Selection of FOP Herbicide Resistant Calli sterility . [0153 ] Callus production from the isolation of immature [0158 ] For sequencing experiments , DNA was isolated embryos (IEs ) was accomplished by removing immature from the both resistant calli and the regenerated plants . The seeds from greenhouse grown sorghum (Inbred BTX430 ) entire 1863 bp Carboxyl Domain (CT ) region was PCR panicles approximately 14 days after anthesis and surface amplified using the four pairs of the overlapping primers sterilized in ethanol followed by a bleach solution and then ( Table 1 ) . washed in sterilized water. Immature embryos were isolated from seeds and transferred to medium with 20 IEs per plate . TABLE 1 Cultures were grown for two weeks at ( 28° C . ) . All explants Primers used to amplify the CT domain region which produced somatic embryogenic calli were transferred of sorghum ACC qene . to growth medium and cultured for an additional 4 - 6 days at PCR Product SEQ 28° C . Explants were transferred to fresh media every three pri Size ID weeks until the entire callus turned into compact and friable mers DNA sequences ( bp ) NO : green callus. Three weeks old freshly subcultured calli were 1F 5 ! GCAACTCTGGTGCTAGGATTGGCA3 ! 553 11 used for the experiment 1R 5 ! GAACATAGCTGAGCCACCTCAATATATT3 ' 12 [ 0154 ] The herbicide active ingredient molecule of ACC herbicide Assure II ( Quizalofop - p - ethyl ) was used for 5 ' GGTGGTCCTAAGATCATGGCGACC3 ' 790 w screening and selection process . For identifying the opti NN 5 ' AGTCTTGGAGTTCCTCTGACCTGAAC3 ' A mum concentration for screening , a kill curve experiment 5 ' CAGCTTGATTCCCATGAGCGATC35 406406 15 with concentrations ranging from 0 .05 to 1 . 0 UM of a . i . ?? 5 ' CCATACAGTCTTGGAGTTCCTCTGA3 ' 16 molecules were tested . Three week old calli were cut into small pieces of 4 - 5 mm dia and 25 pieces were tested in three 5 ' GAGTGTTATGCTGAGAGGACTGCCAA3 ' 711 17 replications for each concentration . Concentrations used 10 5 ' ACCAAGGACCTTCTTGACTTCCTG3 ' 18 were : 0 . 05 , 0 . 1 , 0 . 2 , 0 . 3 , 0 . 4 , 0 . 5 and 1 uM Quizalofop p - ethyl. As 50 % of calli were killed at 0 . 2 uM concentration , [0159 ] Two step PCR reaction was performed using New the following selection criteria was followed for entire England Biolabs multiplex PCR master mix kit , in a 50 ul screening process . Selection process was started with 0 .05 reaction mixture containing 2 ul of total DNA ( 100 ng) , 10 um and doubled the concentration for every 3 weeks and the ul 5xPCR buffer, 2 ul of primer mix ( 5 um / ul of each calli surviving on 1 . 0 uM was considered as resistant calli. forward and reverse primer ) , 0 . 7 ul of 10 uM dNTPs , 0 . 5 ul [ 0155 ] For the chemicalmutagenesis , two chemical muta of Phusion high fidelity DNA polymerase and 34 . 8 ul of gens Ethyl methanesulfonate ( EMS) ( 0 . 25 and 0 . 5 % ) and sterile distilled water . The CT domain sequences were Sodium azide ( 0 . 5 and 1 . 0 uM ) were used . These mutagens amplified with the following temperature conditions: Pre at specified concentration were directly added into the incubation at 98° C . for 3 min , then 35 cycles of denaturation culture media . The calli pieces were first cultured on the at 98° C . for 20 sec , annealing and extension at 72° C . for media containing the mutagens for 6 and 12 hrs and then 1 . 2 min , followed by a final extension at 72° C . for 10 min . transferred to the above mentioned selection procedure . A For sequencing reaction , the entire 1 . 863 kb CT domain total of 16 experiments were conducted with varying amount region was PCR amplified using 1F and 4R primers. The of calli pieces. After 21 to 24 weeks of selection , the PCR products were purified using Quiagen columns and surviving calli were considered as herbicide resistant calli. sequenced at University of Chicago Sequencing Facility The surviving resistant calli were further tested on 1, 2, 5 , using the combination of primers mentioned above . The 10 , 20 , 50 and 100 uM concentration of aimolecule Quizalo sequencing results were aligned and compared using Vector fop - p - ethyl and the herbicide Assure II (FIG . 1 ) . NTI program . [0156 ] A total of 19 resistant calli ( FP - 1 to FP - 19 ) were [0160 ] The DNA and amino acid sequences of wild type obtained from 16 experiments . Out of 19 resistant calli , 3 genotype (BTX430 ) were shown in SEQ ID NO : 1 and 6 , resistant calli ( FP - 8 , FP - 11 , and FP - 12 ) were obtained from respectively ( FIG . 4 ) . The sequencing result showed that the chemical mutagenesis . The resistant calli FP - 7 to FP16 were DNA isolated from FP - 7 , FP - 8 , FP - 9 , FP - 10 , FP - 13 , FP - 14 , showed high level of resistant to both Quizalofop -p - ethyl a . i. FP - 15 and , FP - 16 resistant calli or plants have mutation of TGG to TGC ( SEQ ID NO : 2 ) that leads to replacing the molecules and the herbicide Assure II and survived up to 100 amino acid Tryptophan with Cysteine at the amino acid uM concentration ( FIG . 2 ) . codon position aligning with 1999Trp of the blackgrass Example 2 : Plant Regeneration , DNA Isolation and weed A . myosuroides (W1999C ) ACC protein (SEQ ID NO : Sequence Analysis 7 ) . DNA from FP - 11 and FP - 12 resistant calli or plants have mutation of TGG to TCG (SEQ ID NO : 3 ) that leads to [0157 ] The resistant calli were transferred onto regenera replacing Tryptophan with Serine at codon position tion medium either supplemented with 0 or 1 . 0 uM of W1999S , (SEQ ID NO : 8 ) . DNA from FP - 17 has mutation Quizalofop -p -ethyl . Well -developed shoots were transferred of GCA to GTA ( SEQ ID NO : 4 ) that leads to replacing onto hormone free rooting medium containing O or, 1 , 2 . 5 Alanine with Valine at codon position A2004V (SEO ID NO : and 5 UM Quizalofop -p -ethyl . Rooted plants were trans 9 ) . DNA from FP -4 , FP - 5 , FP -6 , FP - 18 and FP - 19 have ferred into the greenhouse and established into a whole plant mutation of TGG to TCG (SEQ ID NO : 5 ) that leads to (FIG . 3 ). Several hundred plants were regenerated from replacing Tryptophan with Serine at codon position US 2018 /0346920 A1 Dec. 6 , 2018

W2027S , (SEQ ID NO : 10 ). Part of the protein mutation application . The herbicide application results showed that sequences were aligned , compared and shown in FIG . 5 . All W1999C mutant sorghum plants were unaffected by herbi the SNP mutations except FP -8 and FP - 11 , showed het cide application even at 4x rate whereas the control plants erozygous conditions for ACC mutation while FP - 8 and were killed at the 1x rate (FIG . 9 ) . The herbicide resistant FP - 11 were homozygous mutants . plants grew into normal plants and set seed . Example 3 : Screening of Herbicide Resistance [0164 ] The resistant plants were grown to maturity and set Under Greenhouse Conditions with Quizalofop seed either by self or cross pollinating with pollen from other Herbicide Assure II sorghum inbred lines. The harvested F , seed was planted in [0161 ] ACC herbicide resistance was tested under green the greenhouse and inheritance of herbicide resistance was house conditions at different growth stages . At first , two to also demonstrated in two week - old young plants by spraying three week old greenhouse grown F . plants generated from Assure II at 2x rate (FIG . 10 ) . Good seed set was noticed in resistant calli were screened along with control plants for most of the plants (FIG . 11) and seed was produced for all herbicide resistance by spraying with 2 . 5 , 5 . 0 , or 10 oz of mutants except A2004V . Assure II herbicide per acre . These rates represent 0 . 5 , 1 . 0 , and 2 . 0 times ( X ) , respectively , the labeled rate for control Example 4 : Development of KASP DNA Markers of wild sorghum or shattercane . Herbicide were applied at for ACC Gene Mutations the field application rate of 15 gallons of herbicide and water mixture per acre either using small hand sprayer or CO2 [0165 ] Four SNP (Single Nucleotide Polymorphisms) pressurized sprayer. Plants were rated 15 days after herbi mutations ( TGG to TGC at codon position 1999 , TGG to cide application as alive or dead . Herbicide application TCG at codon position 1999 , GCA to GTA at codon position results showed that plants regenerated from FP - 4 to and 2004 and TGG to TCG at codon position 2027 ) in the CT FP - 19 were resistant to either 0 . 5x , 1x or 2x rate of herbicide domain of the ACC gene of wild - type sorghum ( SEQ ID application . However plants from FP -7 to FP - 16 were highly NO : 1 ) are responsible for the herbicide resistance in resistant to herbicide application up to a 2x rate ( FIG . 6 ) BTX430 sorghum . The KASP (Kompetitive Allele Specific while FP - 4 , FP -5 , FP6 , FP18 and FP19 showed moderate PCR ) DNA markers were developed for all SNPs except level of resistance up to 1x rate and were dead at 2x rate . mutation at codon 2004 using the following primers ( Table FP - 1 , FP - 2 and FP - 3 were dead after spraying with 1x rate . 2 ) and the procedures described at the LGC web side All the control plants were dead even at 0 . 5x rate . The describing genotyping chemistry using KASP . surviving plants were transferred into bigger pots for further analysis and allowed to set seed . [0166 ] KASP was performed by a real time PCR allelic [0162 ] ACC herbicide resistance was tested under green discrimination assay using a Roche Light Cycler 480 II house conditions at different growth stages. At first, two to thermocycler (Roche Diagnostics GmbH , Roche Applied three week old greenhouse grown F , plants generated from Science , 68298 Mannheim , Germany ) . A PCR reaction mix resistant calli were screened along with control plants for was prepared in a final volume of 10 ul reaction comprising herbicide resistance by spraying with 2 . 5 , 5 . 0 , or 10 oz of of 5 ul of 2xKASP master mix ( LGC ) , 0 . 14 ul primer mix Assure II herbicide per acre . These rates represent 0 . 5 , 1 . 0 , (X + Y + C ), and 25 ng of genomic DNA with DNase - free and 2 . 0 times ( X ), respectively , the labeled rate for control water to make up the final volume. PCR amplification of wild sorghum or shattercane . Herbicide were applied at conditions: one initial denaturation cycle at 94° C . for 15 the field application rate of 15 gallons ofherbicide and water min , followed by 10 cycles of initial amplification at 94° C . mixture per acre either using small hand sprayer or CO , for 20 seconds and annealing / extension at 61 -55° C . for 1 pressurized sprayer . Plants were rated 15 days after herbi min (dropping 0 .6° C . per cycle ) . Then 30 cycles of ampli cide application as alive or dead . Herbicide application fication program at 94° C . for 20 seconds and 55° C . for 1 results showed that plants regenerated from FP - 4 to and min . The result of the allelic discrimination was determined FP - 19 were resistant to either 0 .5x , 1x or 2x rate of herbicide by Endpoint detection of fluorescence following Roche application .However plants from FP -7 to FP -16 were highly LightCycler @ 480 Instrument Operator ' s Manual . The resistant to herbicide application up to a 2x rate ( FIG . 6 and KASP assay differentiated the homozygous and heterozy FIG . 7 ) while FP - 4 , FP - 5 , FP6 , FP18 and FP19 showed gous nature of the herbicide resistance and separated the moderate level of resistance up to 1x rate and were dead at wild types in segregating populations (FIG . 12 ) . The devel 2x rate (FIG . 8 ). FP - 1, FP - 2 and FP - 3 were dead after oped KASP markers were used to genotype the resistant spraying with 1x rate. All the control plants were dead even calli, Fo, F , and F2 segregating population . at 0 .5x rate . The surviving plants were transferred into bigger pots for further analysis and allowed to set seed . TABLE 2 [0163 ] Mature Fo sorghum plants were also screened for Primers used in KSAP assay for detecting herbicide resistance by spraying higher field rate of herbi W1999C , W1999S and W2027S mutations . cide Assure II at 8 . 0 , 16 . 0 or 32 oz of Assure II herbicide per acre . These rates represent 1 . 0 , 2 . 0 and 4 . 0 times ( X ) , SEQ PCR ID respectively , the labeled rate for control of wild sorghum or primers shattercane . Six to eight weeks old control (BTX430 ) and SNPs DNA sequences NO : TGG to X (HAX 5 ' GGGCTGGACAAGTGTGG 3 ! 19 W1999C mutant plants were sprayed with 1x , 2x and 4x TGC at dye ) rates using a small hand sprayer at the application rate of 15 1999 Y ( FAM 5 ' GGGCTGGACAAGTGTGC 30 20 gallons of herbicide and water per acre . Each pot containing dye ) 3 to 4 plants were sprayed with 75 to 100 ml of herbicide 5 ' CTGAGCTGTCTTGGTTGCAG 3 ' 21 solution or until runoff . Plants were graded 3 weeks after US 2018 /0346920 A1 Dec . 6 , 2018 19

TABLE 2 - continued pressurized sprayer with spray volume of 15 gallons of herbicide and water mixture per acre . Wild type BTX430 Primers used in KSAP assay for detecting plants generated through tissue culture was used as control. W1999C , W1999S and W2027S mutations . Two weeks after spraying , the plants were rated for resis SEQ tance as alive or dead . Genotyping data of F2 plants showed PCR ID that all the plants from Fi homozygous lines were homozy SNPs primers DNA sequences NO : gous ( TGC ) for herbicide resistance . Except 4 plants , all the F , homozygous plants were alive and exhibited high level of TGG to X ( HAX 5 ' TTGCAGAATCTGGGAACC 3 ! TCG at dye ) herbicide resistance ( Table 3 ) . As expected , the plants from 1999 Y ( FAM 5 ' TTGCAGAATCTGGGAACG3 ' 23 F heterozygous lines showed three kinds of genotypes , dye ) TGC (homozygous mutant ); TGG / C (heterozygous mutant 5 ' GGTCAGCTTGATTCCCATGA3 : 24 for herbicide resistance ) and TGG (wild type ) confirming TGG to X ( HAX 5 ' GTCCACCAGAGAAACCTCTCC3 ' 25 the Mendelian segregation ratio of 1 : 2 : 1 with the Chi square TGC at dye ) value of 0 . 88 , 0 . 24 , 0 .53 and 0 .22 which is less than the table 2027 Y ( FAM 5 ' GTCCACCAGAGAAACCTCTCG3 ' 26 value of 2 . 706 . This means the segregation ratios are in dye ) goodness of fit with no significant difference between 5 ' CGTGAAGGATTGCCTCTGTT3 ' 27 expected and observed value . Herbicide application resulted X : Wild type allele ; Y : Mutant allele ; C : Reverse primer the death of 87 % of wild type plants . The remaining wild type plants did not die , but developed dead heart symptoms. Some of the small heterozygous plants showed slight yel Example 5 . Correlating the Herbicide Resistance lowing phenotypes , but later recovered . The presence of with W1999C Mutation in F , Segregating and W1999C mutation in the F2 segregating population either in Homozygous Lines of BTX430 homozygous ( TGC ) or heterozygous ( TGG / C ) conditions [ 0167 ] The BTX430 sorghum plants with W1999C muta clearly correlated with herbicide resistance with correlation tion showing high level of herbicide resistance were self co efficient values of 0 . 937 , 0 . 841 , 0 .783 , and 0 .715 for pollinated and also cross pollinated with tissue culture F2- P3 , F2 - P20 , F2 - P30 and F2 - P50 , respectively . derived wild type BTX430 to produce subsequent genera [0168 ] In addition , Eighteen plants from F2 homozygous tions . Several lines were grown in the greenhouse to matu W1999C mutants line (P12 ) along with control BTX430 rity to set seed . Except few plants , most of the Fo plants were plants were tested for herbicide resistance by spraying with fertile and shed pollen . Plants regenerated from FP - 7 , FP - 9 , 2x and 4x rates ( 16 and 32 ozlacre ) using CO , pressurized FP - 10 , FP - 13 , FP - 14 , FP - 15 and FP - 16 resistant calli all sprayer with spray volume of 15 gallons of herbicide and produced F seed in the greenhouse. Twenty F , seed for each water mixture per acre . Plants were rated 14 days after line were planted in the greenhouse and screened with the herbicide application as alive and dead . The herbicide appli application of herbicide at the 2x ( 16 . 0 oz/ acre ) herbicide cation results showed that all the homozygous F2 plants rate. A total of 54 surviving F? plants (F2 - P1 to F , - P54 ) survived both at 2x ( FIGS . 13A and 13B ) and 4x ( FIGS . comprising of 22 homozygous and 32 heterozygous plants 14A and 14B ) rates , and all the control BTX430 plants were were transferred to bigger pots and grown to set seed by self killed at both application rates. TABLE 3 Genotype and phenotype data of selected F , lines . F? genotype for No. of W1999C plants F , genotype F , phenotype Lines mutant tested TGC TG ( G / C ) TGG Alive Dead F > - P8 Homozygous SO F2 -P12 Homozygous 50 F2- P16 Homozygous 50 F , -P29 Homozygous 50 47 F2 -P3 Heterozygous 10 F > - P20 Heterozygous 12 37 F2 - P30 Heterozygous F2 - 50 Heterozygous BTX430 Wild type 0 ooowoom50 pollination . Five plants from each group were also Example 6 . Introgression of Herbicide Resistant sequenced to confirm the mutation and the zygosity . After Trait into Elite Germplasms Through Recurrent harvesting F2 seed upon maturity , four lines from each group Back Crossing (F2 - P8 , F1 - P12 , F2 - P16 and F _ -29 for homozygous ) and ( F - P3, F - P20 , F - P30 and F - P50 ) were [0169 ] Herbicide resistance was introgressed into 33 selected for genotyping and to know the inheritance of inbred lines which includes both R and B lines for sorghum herbicide tolerance . Fifty F2 plants were genotyped and hybrid production . For backcrossing , the elite inbred lines tested for herbicide resistance by spraying Assure II at the 2x were emasculated and crossed with the herbicide resistant rate ( 16 oz/ acre ) at 2 weeks after planting using CO2 BTX430 -CHR - ACC plants as male parent. After pollina US 2018 /0346920 A1 Dec. 6 , 2018 20

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SEQUENCE LISTING

< 160 > NUMBER OF SEQ ID NOS : 27 < 210 > SEQ ID NO 1 < 211 > LENGTH : 1863 < 212 > TYPE : DNA < 213 > ORGANISM : Sorghum 220 > FEATURE : < 221 > NAME / KEY : misc _ feature < 223 > OTHER INFORMATION : ACC Carboxyl Transferase domain sequence (Wild Type BTX430 ) < 400 > SEQUENCE : 1 gcaaactctg gtgctaggat tggcatagct gatgaagtaa aatcttgctt ccgtgttggg 60 US 2018 /0346920 A1 Dec . 6 , 2018 22

- continued tggtctgacg aaggcagccc tgagcgaggg tttcagtaca tctatctgac tgaagaagac 120 tatgcccgta ttagctcttc tgttatagca cataagctgc agctagatag cggtgaaatt 180 aggtggatta ttgactctgt tgtgggcaag gaggatgggc ttggtgttga gaacatacat 240 ggaagtgctg ctatcgccag tgcttattct agggcatatg aggagacatt tacacttaca 300 tttgtgaccg gacggactgt aggaatagga gottatcttg ctagacttgg tatacggtgc 360 atacagcgtc ttgaccagcc gattatttta acagggtttt ctgccctgaa caagctcctt 420 gggcgggaag tgtacagctc ccacatgcag cttggtggtc ctaagatcat ggcgaccaat 480 ggtgttgtcc acctgactgt tccagatgac cttgaaggtg tttccaatat attgaggtgg 540 ctcagctatg ttcctgcaaa cattggtgga cctcttccta ttaccaaacc tttggaccct 600 ccagacagac ctgttgcata catccctgag aacacatgcg atccacgtgc agccatccgt 660 ggtgtagatg acagccaagg gaaatggttg ggtggtatgt ttgacaaaga cagctttgtg 720 gagacatttg aaggatgggc aaaaacagtg gttactggca gagcaaagct tggaggaatt 780 cctgtgggtg tcatagctgt ggagacacag accatgatgc agcttgtccc tgctgatcca 840 ggtcagcttg attcccatga gcgatccgtt cctcgggctg gacaagtgtg gttcccagat 900 tctgcaacca agacagctca ggcattatta gacttcaacc gtgaaggatt gcctctgttt 960 atcctggcta actggagagg tttctctggt ggacagagag atctctttga aggaattctt 1020 caggctgggt caacaattgt cgagaacctt aggacatata atcagcctgc gtttgtctac 1080 attcctatgg ctggagagct tcgtggagga gottgggttg tggtcgatag caaaataaat 1140 ccagaccgca ttgagtgtta tgctgagagg actgccaaag gtaatgttct cgaacctcaa 1200 gggttaattg aaatcaagtt caggtcagag gaactccaag actgtatggg taggcttgac 1260 cccgagttga taaatctgaa agcaaaactc caagatgtaa agcatggaaa tggaagtcta 1320 ccagacatag aatcccttca gaagagtata gaagcacgta cgaaacagtt gctgccttta 1380 tatacccaga ttgcaatacg gtttgctgaa ttgcatgata cttccctaag aatggcagct 1440 aaaggcgtga ttaagaaagt tgtagactgg gaagaatcac gctctttctt ctataaaagg 1500 ctacggagaa ggatctctga agatgttctt gcaaaagaaa taagacatat agtcggtgac 1560 aacttcactc accaatcago aatggagctcatcaaggaat ggtacctggc ttctccagcc 1620 acagcaggaa gcactggatg ggatgacgat gatgcatttg ttgcctggaa ggacagtcct 1680 gaaaactaca atggatatat ccaagagcta agggctcaaa aagtgtctca gtcgctctct 1740 gatctcactg actccagttc agatctacaa gcattctcgc agggtctttc tacgctatta 1800 gataagatgg atccctctca aagagcgaag tttgttcagg aagtcaagaa ggtccttggt 1860 tga 1863

< 210 > SEQ ID NO 2 < 211 > LENGTH : 1863 < 212 > TYPE : DNA < 213 > ORGANISM : Artificial Sequence FEATURE : < 223 > OTHER INFORMATION : Synthetic Polynucleotide < 220 > FEATURE : < 221 > NAME / KEY : misc _ feature < 223 > OTHER INFORMATION : ACC1 ( TGG to TGC ) < 400 > SEQUENCE : 2 US 2018 /0346920 A1 Dec . 6 , 2018 23

- continued gcaaactctg gtgctaggat tggcatagct gatgaagtaa aatcttgctt ccgtgttggg 60 tggtctgacg aaggcagccc tgagcgaggg tttcagtaca tctatctgac tgaagaagac 120 tatgcccgta ttagctcttc tgttatagca cataagctgc agctagatag cggtgaaatt 180 aggtggatta ttgactctgt tgtgggcaag gaggatgggc ttggtgttga gaacatacat 240 ggaagtgctg ctatcgccag tgottattct agggcatatg aggagacatt tacacttaca 300 tttgtgaccg gacggactgt aggaatagga gottatcttg ctagacttgg tatacggtgc 360 atacagcgtc ttgaccagcc aattatttta acagggtttt ctgccctgaa caagctcctt 420 gggcgggaag tgtacagctc ccacatgcag cttggtggtc ctaagatcat ggcgaccaat 480 ggtgttgtcc acctgactgt tccagatgac cttgaaggtg tttccaatat attgaggtgg 540 ctcagctatg ttcctgcaaa cattggtgga cctcttccta ttaccaaacc tttggaccct 600 ccagacagac ctgttgcata catccctgag aacacatgcg atccacgtgc agccatccgt 660 ggtgtagatg acagccaagg gaaatggttg ggtggtatgt ttgacaaaga cagctttgtg 720 gagacatttg aaggatgggc aaaaacagtg gttactggca gagcaaagct tggaggaatt 780 cctgtgggtg tcatagctgt ggagacacag accatgatgc agcttgtccc tgctgatcca 840 ggtcagcttg attcccatga gcgatccgtt cctcgggctg gacaagtgtg cttcccagat 900 tctgcaacca agacagctca ggcattatta gacttcaacc gtgaaggatt gcctctgttt 960 atcctggcta actggagagg tttctctggt ggacagagag atctctttga aggaattctt 1020 caggctgggt caacaattgt gagaacctt aggacatata atcagcctgc gtttgtctac 1080 attcctatgg ctggagagct tcgtggagga gottgggttg tggtcgatag caaaataaat 1140 ccagaccgca ttgagtgtta tgctgagagg actgccaaag gtaatgttct cgaacctcaa 1200 gggttaattg aaatcaagtt caggtcagag gaactccaag actgtatggg taggcttgac 1260 cccgagttga taaatctgaa agcaaaactc caagatgtaa agcatggaaa tggaagtcta 1320 ccagacatag aatcccttca gaagagtata gaagcacgta cgaaacagtt gctgccttta 1380 tatacccaga ttgcaatacg gtttgctgaa ttgcatgata cttccctaag aatggcagct 1440 aaaggcgtga ttaagaaagt tgtagactgg gaagaatcac gctctttctt ctataaaagg 1500 ctacggagaa ggatctctga agatgttctt gcaaaagaaa taagacatat agtcggtgac 1560 aacttcactc accaatcagc aatggagctc atcaaggaat ggtacctggc ttctccagcc 1620 acagcaggaa gcactggatg ggatgacgat gatgcatttg ttgcctggaa ggacagtcct 1680 gaaaactaca atggatatat ccaagagcta agggctcaaa aagtgtctca gtcgctctct 1740 gatctcactg actccagttc agatctacaa gcattctcgc agggtctttc tacgctatta 1800 gataagatgg atccctctca aagagcgaag tttgttcagg aagtcaagaa ggtccttggt 1860 tga 1863

< 210 > SEQ ID NO 3 < 211 > LENGTH : 1863 < 212 > TYPE : DNA < 213 > ORGANISM : Artificial Sequence < 220 > FEATURE : 23 > OTHER INFORMATION : Synthetic Polynucleotide < 220 > FEATURE : < 221 > NAME / KEY : misc _ feature 223 > OTHER INFORMATION : ACC2 ( TGG to TCG ) < 400 > SEQUENCE : 3 US 2018 /0346920 A1 Dec . 6 , 2018 24

- continued gcaaactctg gtgctaggat tggcatagct gatgaagtaa aatcttgctt ccgtgttggg 60 tggtctgacg aaggcagccc tgagcgaggg tttcagtaca tctatctgac tgaagaagac 120 tatgcccgta ttagctcttc tgttatagca cataagctgc agctagatag cggtgaaatt 180 aggtggatta ttgactctgt tgtgggcaag gaggatgggc ttggtgttga gaacatacat 240 ggaagtgctg ctatcgccag tgottattct agggcatatg aggagacatt tacacttaca 300 tttgtgaccg gacggactgt aggaatagga gottatcttg ctagacttgg tatacggtgc 360 atacagcgtc ttgaccagcc aattatttta acagggtttt ctgccctgaa caagctcctt 420 gggcgggaag tgtacagctc ccacatgcag cttggtggtc ctaagatcat ggcgaccaat 480 ggtgttgtcc acctgactgt tocagatgac cttgaaggtg tttccaatat attgaggtgg 540 ctcagctatg ttcctgcaaa cattggtgga cctcttccta ttaccaaacc tttggaccct 600 ccagacagac ctgttgcata catccctgag aacacatgcg atccacgtgc agccatccgt 660 ggtgtagatg acagccaagg gaaatggttg ggtggtatgt ttgacaaaga cagctttgtg 720 gagacatttg aaggatgggc aaaaacagtg gttactggca gagcaaagct tggaggaatt 780 cctgtgggtg tcatagctgt ggagacacag accatgatgc agcttgtccc tgctgatcca 840 ggtcagcttg attcccatga gcgatccgtt cctcgggctg gacaagtgtc gttcccagat 900 tctgcaacca agacagctca ggcattatta gacttcaacc gtgaaggatt gcctctgttt 960 atcctggcta actggagagg tttctctggt ggacagagag atctctttga aggaattctt 1020 caggctgggt caacaattgt gagaacctt aggacatata atcagcctgc gtttgtctac 1080 attcctatgg ctggagagct tcgtggagga gottgggttg tggtcgatag caaaataaat 1140 ccagaccgca ttgagtgtta tgctgagagg actgccaaag gtaatgttct cgaacctcaa 1200 gggttaattg aaatcaagtt caggtcagag gaactccaag actgtatggg taggcttgac 1260 cccgagttga taaatctgaa agcaaaactc caagatgtaa agcatggaaa tggaagtcta 1320 ccagacatag aatcccttca gaagagtata gaagcacgta cgaaacagtt gctgccttta 1380 tatacccaga ttgcaatacg gtttgctgaa ttgcatgata cttccctaag aatggcagct 1440 aaaggcgtga ttaagaaagt tgtagactgg gaagaatcac gctctttctt ctataaaagg 1500 ctacggagaa ggatctctga agatgttctt gcaaaagaaa taagacatat agtcggtgac 1560 aacttcactc accaatcagc aatggagctc atcaaggaat ggtacctggc ttctccagcc 1620 acagcaggaa gcactggatg ggatgacgat gatgcatttg ttgcctggaa ggacagtcct 1680 gaaaactaca atggatatat ccaagagcta agggctcaaa aagtgtctca gtcgctctct 1740 gatctcactg actccagttc agatctacaa gcattctcgc agggtctttc tacgctatta 1800 gataagatgg atccctctca aagagcgaag tttgttcagg aagtcaagaa ggtccttggt 1860 tga 1863

< 210 > SEQ ID NO 4 < 211 > LENGTH : 1863

< 212P > TYPE : DNA < 213 > ORGANISM : Artificial Sequence < 220 > FEATURE : < 223 > OTHER INFORMATION : Synthetic Polynucleotide < 220 > FEATURE : < 221 > NAME / KEY : misc _ feature 223 > OTHER INFORMATION : ACC3 (GCA to GTA ) US 2018 /0346920 A1 Dec . 6 , 2018 25

- continued < 400 > SEQUENCE : 4 gcaaactctg gtgctaggat tggcatagct gatgaagtaa aatcttgctt ccgtgttggg 60 tggtctgacg aaggcagccc tgagcgaggg tttcagtaca tctatctgac tgaagaagac 120 tatgcccgta ttagctcttc tgttatagca cataagctgc agctagatag cggtgaaatt 180 aggtggatta ttgactctgt tgtgggcaag gaggatgggc ttggtgttga gaacatacat 240 ggaagtgctg ctatcgccag tgottattct agggcatatg aggagacatt tacacttaca 300 tttgtgaccg gacggactgt aggaatagga gottatcttg ctagacttgg tatacggtgc 360 atacagcgtc ttgaccagcc aattatttta acagggtttt ctgccctgaa caagctcctt 420 gggcgggaag tgtacagctc ccacatgcag cttggtggtc ctaagatcat ggcgaccaat 480 ggtgttgtcc acctgactgt tocagatgac cttgaaggtg tttccaatat attgaggtgg 540 ctcagctatg ttcctgcaaa cattggtgga cctcttccta ttaccaaacc tttggaccct 600 ccagacagac ctgttgcata catccctgag aacacatgcg atccacgtgc agccatccgt 660 ggtgtagatg acagccaagg gaaatggttg ggtggtatgt ttgacaaaga cagctttgtg 720 gagacatttg aaggatgggc aaaaacagtggttactggca gagcaaagct tggagga att 780 cctgtgggtg tcatagctgt ggagacacag accatgatgc agcttgtccc tgctgatcca 840 ggtcagcttg attcccatga gcgatccgtt cctcgggctg gacaagtgtc gttcccagat 900 tctgtaacca agacagctca ggcattatta gacttcaacc gtgaaggatt gcctctgttt 960 atcctggcta actggagagg tttctctggt ggacagagag atctctttga aggaattctt 1020 caggctgggt caacaattgt cgagaacctt aggacatata atcagcctgc gtttgtctac 1080 attcctatgg ctggagagct tcgtggagga gottgggttg tggtcgatag caaaataaat 1140 ccagaccgca ttgagtgtta tgctgagagg actgccaaag gtaatgttct cgaacctcaa 1200 gggttaattg aaatcaagtt caggt cagag gaactccaag actgtatggg taggcttgac 1260 cccgagttga taaatctgaa agcaaaactc caagatgtaa agcatggaaa tggaagtcta 1320 ccagacatag aatcccttca gaagagtata gaagcacgta cgaaacagtt gctgccttta 1380 tatacccaga ttgcaatacg gtttgctgaa ttgcatgata cttccctaag aatggcagct 1440 aaaggcgtga ttaagaaagt tgtagactgg gaagaatcac gctctttctt ctataaaagg 1500 ctacggagaa ggatctctga agatgttctt gcaaaagaaa taagacatat agtcggtgac 1560 aacttcactc accaatcagc aatggagctc atcaaggaat ggtacctggc ttctccagcc 1620 acagcaggaa gcactggatg ggatgacgat gatgcatttg ttgcctggaa ggacagtcct 1680 gaaaactaca atggatatat ccaagagcta agggctcaaa aagtgtctca gtcgctctct 1740 gatctcactg actccagttc agatctacaa gcattctcgc agggtctttc tacgctatta 1800 gataagatgg atccctctca aagagcgaag tttgttcagg aagtcaagaa ggtccttggt 1860 tga 1863

< 210 > SEQ ID NO 5 < 211 > LENGTH : 1863 < 212E > TYPE : DNA < 213E > ORGANISM : Artificial Sequence FEATURE : < 223 > OTHER INFORMATION : Synthetic Polynucleotide < 220 > FEATURE : 21 > NAME / KEY : misc _ feature < 223 > OTHER INFORMATION : ACC4 ( TGG to TCG ) US 2018 /0346920 A1 Dec . 6 , 2018

- continued

< 400 > SEQUENCE : 5 gcaaactctg gtgctaggat tggcatagct gatgaagtaa aatcttgctt Ccgtgttggg 60 tggtctgacg aaggcagccc tgagcgaggg tttcagtaca tctatctgac tgaagaagac 120 tatgcccgta ttagctcttc tgttatagca cataagctgc agctagatag cggtgaaatt 180 aggtggatta ttgactctgt tgtgggcaag gaggatgggc ttggtgttga gaacatacat 240 ggaagtgctg ctatcgccag tgottattct agggcatatg aggagacatt tacacttaca 300 tttgtgaccg gacggactgt aggaatagga gottatcttg ctagacttgg tatacggtgc 360 atacagcgtc ttgaccagcc aattatttta acagggtttt ctgccctgaa caagctcctt 420 gggcgggaag tgtacagctc ccacatgcag cttggtggtc ctaagatcat ggcgaccaat 480 ggtgttgtcc acctgactgt tccagatgac cttgaaggtg tttccaatat attgaggtgg 540 ctcagctatg ttcctgcaaa cattggtgga cctcttccta ttaccaaacc tttggaccct 600 ccagacagac ctgttgcata catccctgag aacacatgcg atccacgtgc agccatccgt 660 ggtgtagatg acagccaagg gaaatggttg ggtggtatgt ttgacaaaga cagctttgtg 720 gagacatttg aaggatgggc aaaaacagtg gttactggca gagcaaagct tggaggaatt 780 cctgtgggtg tcatagctgt ggagacacag accatgatgc agcttgtccc tgctgatcca 840 ggtcagettg attcccatga gcgatccgtt cctcgggctg gacaagtgtc gttcccagat 900 tctgtaacca agacagctca ggcattatta gacttcaacc gtgaaggatt gcctctgttt 960 atcctggcta actcgagagg tttctctggt ggacagagag atctctttga aggaattctt 1020 caggctgggt caacaattgt cgagaacctt aggacatata atcagcctgc gtttgtctac 1080 attcctatgg ctggagagct tcgtggagga gottgggttg tggtcgatag caaaataaat 1140 ccagaccgca ttgagtgtta tgctgagagg actgccaaag gtaatgttct cgaacctcaa 1200 gggttaattg aaatcaagtt caggtcagag gaactccaag actgtatggg taggottgac 1260 cccgagttga taaatctgaa agcaaaactc caagatgtaa agcatggaaa tggaagtcta 1320 ccagacatag aatcccttca gaagagtata gaagcacgta cgaaacagtt gctgccttta 1380 tatacccaga ttgcaatacg gtttgctgaa ttgcatgata cttccctaag aatggcagct 1440 aaaggcgtga ttaagaaagt tgtagactgg gaagaatcac gctctttctt ctataaaagg 1500 ctacggagaa ggatctctga agatgttctt gcaaaagaaa taagacatat agtcggtgac 1560 aacttcactc accaatcagc aatggagctc atcaaggaat ggtacctggc ttctccagcc 1620 acagcaggaa gcactggatg ggatgacgat gatgcatttg ttgcctggaa ggacagtcct 1680 gaaaactaca atggatatat ccaagagcta agggctcaaa aagtgtctca gtcgctctct 1740 gatctcactg actccagttc agatctacaa gcattctcgc agggtctttc tacgctatta 1800 gataagatgg atccctctca aagagcgaag tttgttcagg aagtcaagaa ggtccttggt 1860 tga 1863

< 210 > SEQ ID NO 6 < 211 > LENGTH : 620 < 212 > TYPE : PRT < 213 > ORGANISM : Sorghum < 220 > FEATURE : < 221 > NAME / KEY : MISC _ FEATURE < 223 > OTHER INFORMATION : Sorghum Wild type CT domain sequence US 2018 /0346920 A1 Dec . 6 , 2018 27

- continued < 400 > SEQUENCE : 6 Ala Asn Ser Gly Ala Arg Ile Gly Ile Ala Asp Glu Val Lys Ser Cys 15 Phe Arg Val Gly Trp Ser Asp Glu Gly Ser Pro Glu Arg Gly Phe Gin Tyr Ile Tyr Leu Thr Glu Glu Asp Tyr Ala Arg Ile Ser Ser Ser Val 45 Ile Ala His Lys Leu Gin Leu Asp Ser Gly Glu Ile Arg Trp Ile Ile Asp Ser Val Val Gly Lys Glu Asp Gly Leu Gly Val Glu Asn Ile His 65 Gly Ser Ala Ala Ile Ala Ser Ala Tyr Ser Arg Ala Tyr Glu Glu Thr 95 Phe Thr Leu Thr Phe Val Thr Gly Arg Thr Val Gly Ile Gly Ala Tyr

Leu Ala Arg Leu Gly Ile Arg Cys Ile Gin Arg Leu Asp Gin Pro Ile 125 Ile Leu Thr Gly Phe Ser Ala Leu Asn Lys Leu Leu Gly Arg Glu Val

Tyr Ser Ser His Met Gin Leu Gly Gly Pro Lys Ile Met Ala Thr Asn 145 Gly Val Val His Leu Thr Val Pro Asp Asp Leu Glu Gly Val Ser Asn 175 Ile Leu Arg Trp Leu Ser Tyr Val Pro Ala Asn Ile Gly Gly Pro Leu

Pro Ile Thr Lys Pro Leu Asp Pro Pro Asp Arg Pro Val Ala Tyr Ile 205 Pro Glu Asn Thr Cys Asp Pro Arg Ala Ala Ile Arg Gly Val Asp Asp

Ser Gin Gly Lys Trp Leu Gly Gly Met Phe Asp Lys Asp Ser Phe Val 225 Glu Thr Phe Glu Gly Trp Ala Lys Thr Val Val Thr Gly Arg Ala Lys 255

y Ile Pro Val Gly Val Ile Ala Val Glu Thr Gin Thr Met

Met Gin Leu Val Pro Ala Asp Pro Gly Gin Leu Asp Ser His Glu Arg 285 Ser Val Pro Arg Ala Gly Gin Val Trp Phe Pro Asp Ser Ala Thr Lys Thr Ala Gin Ala Leu Leu Asp Phe Asn Arg Glu Gly Leu Pro Leu Phe 305 Ile Leu Ala Asn Trp Arg Gly Phe Ser Gly Gly Gin Arg Asp Leu Phe 335 Glu Gly Ile Leu Gin Ala Gly Ser Thr Ile Val Glu Asn Leu Arg Thr

Tyr Asn Gin Pro Ala Phe Val Tyr Ile Pro Met Ala Gly Glu Leu Arg 365 Gly Gly Ala Trp Val Val Val Asp Ser Lys Ile Asn Pro Asp Arg Ile

Glu Cys Tyr Ala Glu Arg Thr Ala Lys Gly Asn Val Leu Glu Pro Gin 385 US 2018 /0346920 A1 Dec . 6 , 2018 28

- continued

Gly Leu Ile Glu Ile Lys Phe Arg Ser Glu Glu Leu Gin Asp Cys Met 415 Gly Arg Leu Asp Pro Glu Leu Ile Asn Leu Lys Ala Lys Leu Gin Asp 425 430 Val Lys His Gly Asn Gly Ser Leu Pro Asp Ile Glu Ser Leu Gin Lys 435 440 445 Ser Ile Glu Ala Arg Thr Lys Gin Leu Leu Pro Leu Tyr Thr Gin Ile

Ala Ile Arq Phe Ala Glu Leu His Asp Thr Ser Leu Arq Met Ala Ala 465 480 Lys Gly Val Ile Lys Lys Val Val Asp Trp Glu Glu Ser Arg Ser Phe 495 Phe Tyr Lys Arg Leu Arg Arg Arg Ile Ser Glu Asp Val Leu Ala Lys 505 510 Glu Ile Arg His Ile Val Gly Asp Asn Phe Thr His Gin Ser Ala Met 515 520 525 Glu Leu Ile Lys Glu Trp Tyr Leu Ala Ser Pro Ala Thr Ala Gly Ser Thr Gly Trp Asp Asp Asp Asp Ala Phe Val Ala Trp Lys Asp Ser Pro 545 560 Glu Asn Tyr Asn Gly Tyr Ile Gin Glu Leu Arg Ala Gin Lys Val Ser 575 Gin Ser Leu Ser Asp Leu Thr Asp Ser Ser Ser Asp Leu Gin Ala Phe 585 590 Ser Gln Gly Leu Ser Thr Leu Leu Asp Lys Met Asp Pro Ser Gln Arq 595 600 605 Ala Lys Phe Val Gln Glu Val Lys Lys Val Leu Gly 610 615 620

< 210 > SEQ ID NO 7 < 211 > LENGTH : 620 < 212 > TYPE : PRT < 213 > ORGANISM : Artificial Sequence < 220 > FEATURE : < 223 > OTHER INFORMATION : Synthetic Polypeptide < 220NNNNNEPP > FEATURE : < 221 > NAME / KEY : MISC FEATURE < 223 > OTHER INFORMATION : ACCI (W1999C ) < 400 > SEQUENCE : 7 Ala Asn Ser Gly Ala Arg Ile Gly Ile Ala Asp Glu Val Lys Ser Cys Phe Arg Val Gly Trp Ser Asp Glu Gly Ser Pro Glu Arg Gly Phe Gin 20 25 Tyr Ile Tyr Leu Thr Glu Glu Asp Tyr Ala Arg Ile Ser Ser Ser Val

Ile Ala His Lys Leu Gln Leu Asp Ser Gly Glu Ile Arq Trp Ile Ile 55Leu Asp Ser Gly Glu Ile Arg Trp Ile Ile Asp Ser Val Val Gly Lys Glu Asp Gly Leu Gly Val Glu Asn Ile His

Gly Ser Ala Ala Ile Ala Ser Ala Tyr Ser Arg Ala Tyr Glu Glu Thr

Phe Thr Leu Thr Phe Val Thr Gly Arg Thr Val Gly Ile Gly Ala Tyr 100 105 US 2018 /0346920 A1 Dec . 6 , 2018

- continued

Leu Ala Arg Leu Gly Ile Arg Cys Ile Gin Arg Leu Asp Gin Pro Ile 115 120 Ile Leu Thr Gly Phe Ser Ala Leu Asn Lys Leu Leu Gly Arg Glu Val 130 135 Tyr Ser Ser His Met Gin Leu Gly Gly Pro Lys Ile Met Ala Thr Asn 145 150 155 160 Gly Val Val His Leu Thr Val Pro Asp Asp Leu Glu Gly Val Ser Asn 170 175 Ile Leu Arg Trp Leu Ser Tyr Val Pro Ala Asn Ile Gly Gly Pro Leu 190 Pro Ile Thr Lys Pro Leu Asp Pro Pro Asp Arg Pro Val Ala Tyr Ile 195 200 Pro Glu Asn Thr Cys Asp Pro Arg Ala Ala Ile Arg Gly Val Asp Asp 210 215 Ser Gin Gly Lys Trp Leu Gly Gly Met Phe Asp Lys Asp Ser Phe Val 225 230 235 240 Glu Thr Phe Glu Gly Trp Ala Lys Thr Val Val Thr Gly Arg Ala Lys 250 255 Leu Gly Gly Ile Pro Val Gly Val Ile Ala Val Glu Thr Gin Thr Met 270 Met Gin Leu Val Pro Ala Asp Pro Gly Gin Leu Asp Ser His Glu Arg 275 280 Ser Val Pro Arg Ala Gly Gin Val Cys Phe Pro Asp Ser Ala Thr Lys 290 295 Thr Ala Gin Ala Leu Leu Asp Phe Asn Arg Glu Gly Leu Pro Leu Phe 305 310 315 320 Ile Leu Ala Asn Trp Arg Gly Phe Ser Gly Gly Gin Arg Asp Leu Phe 330 335

Glu Gly Ile Leu Gln Ala Gly Ser Thr Ile Val Glu Asn Leu Arq Thr 350 Tyr Asn Gin Pro Ala Phe Val Tyr Ile Pro Met Ala G y Glu Leu Arg 355 360 Gly Gly Ala Trp Val Val Val Asp Ser Lys Ile Asn Pro Asp Arg Ile 370 375 Glu Cys Tyr Ala Glu Arg Thr Ala Lys Gly Asn Val Leu Glu Pro Gin 385 390 395 400 Gly Leu Ile Glu Ile Lys Phe Arg Ser Glu Glu Leu Gin Asp Cys Met 410 415 Gly Arg Leu Asp Pro Glu Leu Ile Asn Leu Lys Ala Lys Leu Gin Asp 430 Val Lys His Gly Asn Gly Ser Leu Pro Asp Ile Glu Ser Leu Gin Lys 435 440 Ser Ile Glu Ala Arg Thr Lys Gin Leu Leu Pro Leu Tyr Thr Gin Ile 450 455 Ala Ile Arg Phe Ala Glu Leu His Asp Thr Ser Leu Arg Met Ala Ala 465 470 475 480 Lys Gly Val Ile Lys Lys Val Val Asp Trp Glu Glu Ser Arg Ser Phe 490 495 Phe Tyr Lys Arg Leu Arg Arg Arg Ile Ser Glu Asp Val Leu Ala Lys 510 US 2018 /0346920 A1 Dec . 6 , 2018 30

- continued Glu Ile Arg His Ile Val Gly Asp Asn Phe Thr His Gin Ser Ala Met 515 520 525 Glu Leu Ile Lys Glu Trp Tyr Leu Ala Ser Pro Ala Thr Ala Gly Ser 530 535 540 Thr Gly Trp Asp Asp Asp Asp Ala Phe Val Ala Trp Lys Asp Ser Pro 545 550 555 560 Glu Asn Tyr Asn Gly Tyr Ile Gin Glu Leu Arg Ala Gin Lys Val Ser 570 575 Gin Ser Leu Ser Asp Leu Thr Asp Ser Ser Ser Asp Leu Gin Ala Phe 580 585 590 Ser Gin Gly Leu Ser Thr Leu Leu Asp Lys Met Asp Pro Ser Gin Arg 595 600 605 Ala Lys Phe Val Gln Glu Val Lys Lys Val Leu Gly 610 615 620

< 210 > SEQ ID NO 8 < 211 > LENGTH : 620 < 212 > TYPE : PRT < 213 > ORGANISM : Artificial Sequence < 220 > FEATURE : < 223 > OTHER INFORMATION : Synthetic Polypeptide < 220 > FEATURE : 21 > NAME / KEY : MISC _ FEATURE < 223 > OTHER INFORMATION : ACC2 (W1999S ) < 400 > SEQUENCE : 8 Ala Asn Ser Gly Ala Arg Ile Gly Ile Ala Asp Glu Val Lys Ser Cys 15 Phe Arg Val Gly Trp Ser Asp Glu Gly Ser Pro Glu Arg Gly Phe Gin

Tyr Ile Tyr Leu Thr Glu Glu Asp Tyr Ala Arg Ile Ser Ser Ser Val

Ile Ala His Lys Leu Gln Leu Asp Ser Gly Glu Ile Ara Trp Ile Ile

Asp Ser Val Val Gly Lys Glu Asp Gly Leu Gly Val Glu Asn Ile His Gly Ser Ala Ala Ile Ala Ser Ala Tyr Ser Arg Ala Tyr Glu Glu Thr 95 Phe Thr Leu Thr Phe Val Thr Gly Arg Thr Val Gly Ile Gly Ala Tyr Leu Ala Arg Leu Gly Ile Arg Cys Ile Gin Arg Leu Asp Gin Pro Ile Ile Leu Thr Gly Phe Ser Ala Leu Asn Lys Leu Leu Gly Arg Glu Val Tyr Ser Ser His Met Gin Leu Gly Gly Pro Lys Ile Met Ala Thr Asn

Gly Val Val His Leu Thr Val Pro Asp Asp Leu Glu Gly Val Ser Asn 175 Ile Leu Arg Trp Leu Ser Tyr Val Pro Ala Asn Ile Gly Gly Pro Leu

Pro Ile Thr Lys Pro Leu Asp Pro Pro Asp Arg Pro Val Ala Tyr Ile

Pro Glu Asn Thr Cys Asp Pro Arg Ala Ala Ile Arg Gly Val Asp Asp US 2018 /0346920 A1 Dec . 6 , 2018 31

- continued Ser Gin Gly Lys Trp Leu Gly Gly Met Phe Asp Lys Asp Ser Phe Val 225 230 235 240 Glu Thr Phe Glu Gly Trp Ala Lys Thr Val Val Thr Gly Arg Ala Lys 255 Leu Gly Gly Ile Pro Val Gly Val Ile Ala Val Glu Thr Gin Thr Met 260 265 270 Met Gin Leu Val Pro Ala Asp Pro Gly Gin Leu Asp Ser His Glu Arg 280 Ser Val Pro Arg Ala Gly Gin Val Ser Phe Pro Asp Ser Ala Thr Lys 295 Thr Ala Gln Ala Leu Leu Asp Phe Asn Arg Glu Gly Leu Pro Leu Phe 305 310 315 320 Ile Leu Ala Asn Trp Arg Gly Phe Ser Gly Gly Gin Arg Asp Leu Phe 335 Glu Gly Ile Leu Gin Ala Gly Ser Thr Ile Val Glu Asn Leu Arg Thr 340 345 350 Tyr Asn Gin Pro Ala Phe Val Tyr Ile Pro Met Ala Gly Glu Leu Arg 360 365 Gly Gly Ala Trp Val Val Val Asp Ser Lys Ile Asn Pro Asp Arg Ile 375 Glu Cys Tyr Ala Glu Arg Thr Ala Lys Gly Asn Val Leu Glu Pro Gin 385 390 395 400 Gly Leu Ile Glu Ile Lys Phe Arg Ser Glu Glu Leu Gin Asp Cys Met 415 Gly Arg Leu Asp Pro Glu Leu Ile Asn Leu Lys Ala Lys Leu Gin Asp 420 425 430

Val Lys His Gly Asn Gly Ser Leu Pro Asp Ile Glu Ser Leu Gln Lys 440 445 Ser Ile Glu Ala Arg Thr Lys Gin Leu Leu Pro Leu Tyr Thr Gin Ile 455 Ala Ile Arg Phe Ala Glu Leu His Asp Thr Ser Leu Arg Met Ala Ala 465 470 475 480 Lys Gly Val Ile Lys Lys Val Val Asp Trp Glu Glu Ser Arg Ser Phe 495 Phe Tyr Lys Arg Leu Arg Arg Arg Ile Ser Glu Asp Val Leu Ala Lys 500 505 510 Glu Ile Arq His Ile Val Gly Asp Asn Phe Thr His Gin Ser Ala Met 520 525 Glu Leu Ile Lys Glu Trp Tyr Leu Ala Ser Pro Ala Thr Ala Gly Ser 535 Thr Gly Trp Asp Asp Asp Asp Ala Phe Val Ala Trp Lys Asp Ser Pro 545 550 555 560 Glu Asn Tyr Asn Gly Tyr Ile Gin Glu Leu Arg Ala Gin Lys Val Ser 575

Gln Ser Leu Ser Asp Leu Thr Asp Ser Ser Ser Asp Leu Gln Ala Phe 580 585 590 Ser Gin Gly Leu Ser Thr Leu Leu Asp Lys Met Asp Pro Ser Gin Arg 600 605 Ala Lys Phe Val Gin Glu Val Lys Lys Val Leu Gly 615 US 2018 /0346920 A1 Dec . 6 , 2018 32

- continued < 210 > SEQ ID NO 9 < 211 > LENGTH : 620 < 212 > TYPE : PRT < 213WNHO > ORGANISM : Artificial Sequence < 220 > FEATURE : < 223 > OTHER INFORMATION : Synthetic Polypeptide < 220 > FEATURE : < 221 > NAME / KEY : MISC _ FEATURE < 223 > OTHER INFORMATION : ACC3 ( A2004V ) < 400 > SEQUENCE : 9 Ala Asn Ser Gly Ala Arg Ile Gly Ile Ala Asp Glu Val Lys Ser Cys 15 Phe Arg Val Gly Trp Ser Asp Glu Gly Ser Pro Glu Arg Gly Phe Gin 20 30

Tyr Ile Tyr Leu Thr Glu Glu Asp Tyr Ala Arg Ile Ser Ser Ser Val 45 Ile Ala His Lys Leu Gin Leu Asp Ser Gly Glu Ile Arg Trp Ile Ile Asp Ser Val Val Gly Lys Glu Asp Gly Leu Gly Val Glu Asn Ile His 75 80

Gly Ser Ala Ala Ile Ala Ser Ala Tyr Ser Arg Ala Tyr Glu Glu Thr 95 Phe Thr Leu Thr Phe Val Thr Gly Arg Thr Val Gly Ile Gly Ala Tyr 100 110 Leu Ala Arg Leu Gly Ile Arg Cys Ile Gin Arg Leu Asp Gin Pro Ile 125 Ile Leu Thr Gly Phe Ser Ala Leu Asn Lys Leu Leu Gly Arg Glu Val

Tyr Ser Ser His Met Gln Leu Gly Gly Pro Lys Ile Met Ala Thr Asn 155 160 Gly Val Val His Leu Thr Val Pro Asp Asp Leu Glu Gly Val Ser Asn 175 Ile Leu Arg Trp Leu Ser Tyr Val Pro Ala Asn Ile Gly Gly Pro Leu 180 190 Pro Ile Thr Lys Pro Leu Asp Pro Pro Asp Arg Pro Val Ala Tyr Ile 205 Pro Glu Asn Thr Cys Asp Pro Arg Ala Ala Ile Arg Gly Val Asp Asp

Ser Gin Gly Lys Trp Leu Gly Gly Met Phe Asp Lys Asp Ser Phe Val 235 240 Glu Thr Phe Glu Gly Trp Ala Lys Thr Val Val Thr Gly Arg Ala Lys 255 Leu Gly Gly Ile Pro Val Gly Val Ile Ala Val Glu Thr Gin Thr Met 260 270 Met Gin Leu Val Pro Ala Asp Pro Gly Gin Leu Asp Ser His Glu Arg 285 Ser Val Pro Arg Ala Gly Gin Val Trp Phe Pro Asp Ser Val Thr Lys

Thr Ala Gln Ala Leu Leu Asp Phe Asn Arq Glu Gly Leu Pro Leu Phe 315 320 Ile Leu Ala Asn Trp Arg Gly Phe Ser Gly Gly Gin Arg Asp Leu Phe 335 Glu Gly Ile Leu Gin Ala Gly Ser Thr Ile Val Glu Asn Leu Arg Thr US 2018 /0346920 A1 Dec . 6 , 2018 33

- continued 340 345 350 Tyr Asn Gin Pro Ala Phe Val Tyr Ile Pro Met Ala Gly Glu Leu Arq 360 365 Gly Gly Ala Trp Val Val Val Asp Ser Lys Ile Asn Pro Asp Arg Ile 380 Glu Cys Tyr Ala Glu Arg Thr Ala Lys Gly Asn Val Leu Glu Pro Gin 385 395 400 Gly Leu Ile Glu Ile Lys Phe Arg Ser Glu Glu Leu Gin Asp Cys Met 415 Gly Arg Leu Asp Pro Glu Leu Ile Asn Leu Lys Ala Lys Leu Gin Asp 425 430 Val Lys His Gly Asn Gly Ser Leu Pro Asp Ile Glu Ser Leu Gin Lys 440 445 Ser Ile Glu Ala Arg Thr Lys Gin Leu Leu Pro Leu Tyr Thr Gin Ile 460

Ala Ile Arg Phe Ala Glu Leu His Asp Thr Ser Leu Arg Met Ala Ala 465 475 480 Lys Gly Val Ile Lys Lys Val Val Asp Trp Glu Glu Ser Arg Ser Phe 495 Phe Tyr Lys Arg Leu Arg Arg Arg Ile Ser Glu Asp Val Leu Ala Lys 505 510 Glu Ile Arg His Ile Val Gly Asp Asn Phe Thr His Gin Ser Ala Met 520 525 Glu Leu Ile Lys Glu Trp Tyr Leu Ala Ser Pro Ala Thr Ala Gly Ser 540 Thr Gly Trp Asp Asp Asp Asp Ala Phe Val Ala Trp Lys Asp Ser Pro 545 555 560 Glu Asn Tyr Asn Gly Tyr Ile Gin Glu Leu Arg Ala Gin Lys Val Ser 575 Gin Ser Leu Ser Asp Leu Thr Asp Ser Ser Ser Asp Leu Gin Ala Phe 585 590 Ser Gin Gly Leu Ser Thr Leu Leu Asp Lys Met Asp Pro Ser Gin Arg 600 605 Ala Lys Phe Val Gin Glu Val Lys Lys Val Leu Gly 610 620

< 210 > SEQ ID NO 10 < 211 > LENGTH : 620 < 212NI > TYPE : PRT

< 213OW > ORGANISM : Artificial Sequence < 220 > FEATURE : < 223W > OTHER INFORMATION : Synthetic Polypeptide < 220O > FEATURE : < 221NNNNAH > NAME / KEY : MISC _ FEATURE < 223 > OTHER INFORMATION : ACC4 (W20275 ) < 400 > SEQUENCE : 10 Ala Asn Ser Gly Ala Arg Ile Gly Ile Ala Asp Glu Val Lys Ser Cys 10 15 Phe Arg Val Gly Trp Ser Asp Glu Gly Ser Pro Glu Arg Gly Phe Gin 25 Tyr Ile Tyr Leu Thr Glu Glu Asp Tyr Ala Arg Ile Ser Ser Ser Val 45 Ile Ala His Lys Leu Gin Leu Asp Ser Gly Glu Ile Arg Trp Ile Ile US 2018 /0346920 A1 Dec . 6 , 2018 34

- continued

55 60 Asp Ser Val Val Gly Lys Glu Asp Gly Leu Gly Val Glu Asn Ile His 70 80 Gly Ser Ala Ala Ile Ala Ser Ala Tyr Ser Arg Ala Tyr Glu Glu Thr 95 Phe Thr Leu Thr Phe Val Thr Gly Arg Thr Val Gly Ile Gly Ala Tyr 105 110 Leu Ala Arg Leu Gly Ile Arg Cys Ile Gin Arg Leu Asp Gin Pro Ile 115 120 Ile Leu Thr Gly Phe Ser Ala Leu Asn Lys Leu Leu Gly Arg Glu Val

Tyr Ser Ser His Met Gln Leu Gly Gly Pro Lys Ile Met Ala Thr Asn 150 160 Gly Val Val His Leu Thr Val Pro Asp Asp Leu Glu Gly Val Ser Asn 175 Ile Leu Arg Trp Leu Ser Tyr Val Pro Ala Asn Ile Gly Gly Pro Leu 185 190 Pro Ile Thr Lys Pro Leu Asp Pro Pro Asp Arg Pro Val Ala Tyr Ile 195 200 Pro Glu Asn Thr Cys Asp Pro Arg Ala Ala Ile Arg Gly Val Asp Asp Ser Gin Gly Lys Trp Leu Gly Gly Met Phe Asp Lys Asp Ser Phe Val 230 240 Glu Thr Phe Glu Gly Trp Ala Lys Thr Val Val Thr Gly Arg Ala Lys 255 Leu Gly Gly Ile Pro Val Gly Val Ile Ala Val Glu Thr Gin Thr Met 265 270 Met Gin Leu Val Pro Ala Asp Pro Gly Gin Leu Asp Ser His Glu Arg 275 280 Ser Val Pro Arg Ala Gly Gin Val Trp Phe Pro Asp Ser Ala Thr Lys

Thr Ala Gin Ala Leu Leu Asp Phe Asn Arg Glu Gly Leu Pro Leu Phe 310 320 Ile Leu Ala Asn Ser Arg Gly Phe Ser Gly Gly Gin Arg Asp Leu Phe 335 Glu Gly Ile Leu Gin Ala Gly Ser Thr Ile Val Glu Asn Leu Arg Thr 345 350 Tyr Asn Gin Pro Ala Phe Val Tyr Ile Pro Met Ala Gly Glu Leu Arg 355 360 Gly Gly Ala Trp Val Val Val Asp Ser Lys Ile Asn Pro Asp Arg Ile Glu Cys Tyr Ala Glu Arg Thr Ala Lys Gly Asn Val Leu Glu Pro Gin 390 400 Gly Leu Ile Glu Ile Lys Phe Arg Ser Glu Glu Leu Gin Asp Cys Met 415 Gly Arg Leu Asp Pro Glu Leu Ile Asn Leu Lys Ala Lys Leu Gin Asp 425 430 Val Lys His Gly Asn Gly Ser Leu Pro Asp Ile Glu Ser Leu Gin Lys 435 440 Ser Ile Glu Ala Arg Thr Lys Gin Leu Leu Pro Leu Tyr Thr Gin Ile US 2018 /0346920 A1 Dec . 6 , 2018 35

- continued

Ala Ile Arq Phe Ala Glu Leu His Asp Thr Ser Leu Arq Met Ala Ala 470 475 480 Lys Gly Val Ile Lys Lys Val Val Asp Trp Glu Glu Ser Arg Ser Phe 495 Phe Tyr Lys Arg Leu Arg Arg Arg Ile Ser Glu Asp Val Leu Ala Lys 505 Glu Ile Arg His Ile Val Gly Asp Asn Phe Thr His Gin Ser Ala Met 525 Glu Leu Ile Lys Glu Trp Tyr Leu Ala Ser Pro Ala Thr Ala Gly Ser 540 Thr Gly Trp Asp Asp Asp Asp Ala Phe Val Ala Trp Lys Asp Ser Pro 550 555 560 Glu Asn Tyr Asn Gly Tyr Ile Gin Glu Leu Arg Ala Gin Lys Val Ser 575 Gin Ser Leu Ser Asp Leu Thr Asp Ser Ser Ser Asp Leu Gin Ala Phe 585 Ser Gin Gly Leu Ser Thr Leu Leu Asp Lys Met Asp Pro Ser Gin Arg 605 Ala Lys Phe Val Gln Glu Val Lys Lys Val Leu Gly 615 620

< 210 > SEQ ID NO 11 < 211 > LENGTH : 24 < 212 > TYPE : DNA < 213 > ORGANISM : Artificial Sequence < 220 > FEATURE : < 223 > OTHER INFORMATION : Synthetic Primer < 220 > FEATURE : V 21 > NAME / KEY : misc _ feature < 223 > OTHER INFORMATION : PCR primer 17 < 400 > SEQUENCE : 11 gcaactctgg tgctaggatt ggca 24

< 210 > SEO ID NO 12 < 211 > LENGTH : 28 < 212 > TYPE : DNA < 213 > ORGANISM : Artificial Sequence < 220 > FEATURE : < 223 > OTHER INFORMATION : Synthetic Primer < 220 > FEATURE : < 221 > NAME /KEY : misc _ feature < 223 > OTHER INFORMATION : PCR Primer 1R < 400 > SEQUENCE : 12 gaacatagct gagccacctc aatatatt 28

< 210 > SEQ ID NO 13 < 211 > LENGTH : 24

< 212P > TYPE : DNA < 213 > ORGANISM : Artificial Sequence P

N FEATURE : < 223 > OTHER INFORMATION : Synthetic Primer < 220 > FEATURE : < 221 > NAME / KEY : misc _ feature < 223 > OTHER INFORMATION : PCR Primer 2F < 400 > SEQUENCE : 13 ggtggtccta agatcatggc gacc 24 US 2018 /0346920 A1 Dec . 6 , 2018 36

- continued

< 210 > SEQ ID NO 14 < 211 > LENGTH : 26 < 212 > TYPE : DNA < 213 > ORGANISM : Artificial Sequence < 220 > FEATURE : < 223 > OTHER INFORMATION : Synthetic Primer < 220 > FEATURE : 1 > NAME / KEY : misc _ feature < 223 > OTHER INFORMATION : PCR Primer 2R < 400 > SEQUENCE : 14 agtcttggag ttcctctgac ctgaac 26

< 210 > SEQ ID NO 15 < 211 > LENGTH : 23 < 212 > TYPE : DNA < 213 > ORGANISM : Artificial Sequence < 2 FEATURE : < 223OWNWO > OTHER INFORMATION : Synethic Primer < 220 > FEATURE : < 221 > NAME / KEY : misc _ feature < 223 > OTHER INFORMATION : PCR Primer 3F < 400 > SEQUENCE : 15 cagcttgatt cccatgagcg atc 3

< 210 > SEQ ID NO 16 < 211 > LENGTH : 25 < 212 > TYPE : DNA V< 213 > ORGANISM : Artificial Sequence V< 220 > FEATURE : V< 223 > OTHER INFORMATION : Synthetic Primer V< 220 > FEATURE : V< 221 > NAME / KEY : misc _ feature V< 223NNNNNAH > OTHER INFORMATION : PCR Primer 3R < 400 > SEQUENCE : 16 ccatacagtc ttggagttcc tctga 25

< 210 > SEQ ID NO 17 < 211 > LENGTH : 26 < 212 > TYPE : DNA < 213 > ORGANISM : Artificial Sequence < 220 > FEATURE : < 223 > OTHER INFORMATION : Synthetic Polynucleotide < 400 > SEQUENCE : 17 gagtgttatg ctgagaggac tgccaa 26

< 210 > SEQ ID NO 18 < 211 > LENGTH : 24 < 212 > TYPE : DNA < 213 > ORGANISM : Artificial Sequence < 220 > FEATURE : < 223 > OTHER INFORMATION : Synthetic Primer < 220 > FEATURE : < 221 > NAME / KEY : misc _ feature < 223 > OTHER INFORMATION : PCR Primer 4R < 400 > SEQUENCE : 18 accaaggacc ttcttgactt cctg 24

< 210 > SEQ ID NO 19 < 211 > LENGTH : 17 < 212 > TYPE : DNA US 2018 /0346920 A1 Dec . 6 , 2018 37

- continued < 213 > ORGANISM : Artificial Sequence < 220 > FEATURE : < 223 > OTHER INFORMATION : Synthetic Primer < 220 > FEATURE : < 221 > NAME /KEY : misc _ feature < 223 > OTHER INFORMATION : X ( HAX dye ) < 400 > SEQUENCE : 19 gggctggaca agtgtgg 17

< 210 > SEQ ID NO 20 < 211 > LENGTH : 17 V< 212 > TYPE : DNA V< 213 > ORGANISM : Artificial Sequence V< 220 > FEATURE : V< 223WFOWoin > OTHER INFORMATION : Synthetic Primer V< 220 > FEATURE : V< 221 > NAME /KEY : misc _ feature V< 223 > OTHER INFORMATION : Y ( FAM dye ) < 400 > SEQUENCE : 20 gggctggaca agtgtgc 17

< 210 > SEQ ID NO 21 < 211 > LENGTH : 20 < 212 > TYPE : DNA < 213 > ORGANISM : Artificial Sequence < 220 > FEATURE : < 223 > OTHER INFORMATION : Synthetic Primer < 220 > FEATURE : < 221 > NAME / KEY : misc _ feature < 223 > OTHER INFORMATION : C < 400 > SEQUENCE : 21 ctgagctgtc ttggttgcag 20

< 210 > SEQ ID NO 22 < 211 > LENGTH : 18 < 212 > TYPE : DNA < 213 > ORGANISM : Artificial Sequence < 220 > FEATURE : < 223 > OTHER INFORMATION : Synthetic Primer < 220 > FEATURE : < 221 > NAME / KEY : misc _ feature < 223 > OTHER INFORMATION : X (HAX dye ) < 400 > SEQUENCE : 22 ttgcagaatc tgggaacc 18

< 210 > SEQ ID NO 23 < 211 > LENGTH : 18 < 212 > TYPE : DNA < 213 > ORGANISM : Artificial Sequence < 220 > FEATURE : < 223 > OTHER INFORMATION : Synthetic Primer 220 > FEATURE : 21 > NAME / KEY : misc feature < 223 > OTHER INFORMATION : Y ( FAM dye ) < 400 > SEQUENCE : 23 ttgcagaatc tgggaacg 18

< 210 > SEQ ID NO 24 < 211 > LENGTH : 20 < 212 > TYPE : DNA < 213 > ORGANISM : Artificial Sequence US 2018 /0346920 A1 Dec . 6 , 2018 38

- continued < 220 > FEATURE : < 223 > OTHER INFORMATION : Synthetic Primer < 220 > FEATURE : < 221OM > NAME / KEY : misc _ feature < 223 > OTHER INFORMATION : C < 400 > SEQUENCE : 24 ggtcagettg attcccatga 20

< 210 > SEQ ID NO 25 < 211 > LENGTH : 21 < 212 > TYPE : DNA < 213 > ORGANISM : Artificial Sequence FEATURE : < 223 > OTHER INFORMATION : Synthetic Primer 20 > FEATURE : < 221 > NAME / KEY : misc _ feature < 223 > OTHER INFORMATION : X (HAX dye ) < 400 > SEQUENCE : 25 gtccaccaga gaaacctctcc 21

< 210 > SEQ ID NO 26 < 211 > LENGTH : 21 < 212 > TYPE : DNA < 213 > ORGANISM : Artificial Sequence FEATURE : < 223 > OTHER INFORMATION : Synthetic Primer < 220 > FEATURE : < 221 > NAME / KEY : misc _ feature 23 > OTHER INFORMATION : Y ( FAM dye ) < 400 > SEQUENCE : 26 gtccaccaga gaaacctctc g 21

< 210 > SEQ ID NO 27 < 211 > LENGTH : 20 < 212 > TYPE : DNA < 213 > ORGANISM : Artificial Sequence < 220 > FEATURE : < 223 > OTHER INFORMATION : Synthetic Primer < 220 > FEATURE : < 221 > NAME / KEY : misc _ feature < 223 > OTHER INFORMATION : C < 400 > SEQUENCE : 27 cgtgaaggat tgcctctgtt 20

1 . An ACC inhibitor herbicide tolerant sorghum plant or the nucleotide sequence of SEQ ID NO : 3, SEQ ID NO : plant part thereof comprising one or more mutations of the 4 or SEO ID NO : 5 ; Acetyl - CoA Carboxylase (ACC ) gene , wherein the sorghum f . the nucleotide sequence of SEQ ID NO : 3 and one of plant or plant part has increased resistance to one or more the following: ACC inhibiting herbicide as compared with a wild - type the nucleotide sequence of SEQ ID NO : 4 or SEQ ID sorghum cultivar or plant. NO : 5 ; 2. The sorghum plant or plant part of claim 1, wherein the g . the nucleotide sequence of SEQ ID NO : 4 and SEQ ID AAC gene is a sorghum acetyl- CoA carboxylase gene , NO : 5 ; wherein the nucleotide sequence encoding the CT domain of h . the nucleotide sequence of SEQ ID NO : 2 , SEQ ID NO : the ACC protein comprises one of the following : 3 and SEQ ID NO : 4 ; a . the nucleotide sequence of SEQ ID NO : 2 ; i. the nucleotide sequence of SEQ ID NO : 2 , SEQ ID NO : b . the nucleotide sequence of SEQ ID NO : 3 ; 3 and SEQ ID NO : 5 ; c . the nucleotide sequence of SEQ ID NO : 4 ; j . the nucleotide sequence of SEQ ID NO : 2, SEQ ID NO : d . the nucleotide sequence of SEQ ID NO : 5 ; 4 and SEQ ID NO : 5 ; or e . the nucleotide sequence of SEQ ID NO : 2 and one of k . The nucleotide sequence of SEQ ID NO : 3 , SEQ ID the following: NO : 4 and SEQ ID NO : 5 . US 2018 /0346920 A1 Dec. 6 , 2018 39

3 . The sorghum plant or plantpart of claim 1 , wherein said phan to Serine amino acid substitution at an amino acid AAC gene encodes a sorghum acetyl- CoA protein having a position 1999 (W1999S ) aligning with the amino acid CT domain comprising one or more of the following muta sequence of SEQ ID NO : 6 and a Tryptophan to Serine tions: amino acid substitution at an amino acid position 2027 a . a Tryptophan to Cysteine amino acid substitution at an (W2027S ) aligning with the amino acid sequence of amino acid position 1999 (W1999C ) aligning with the SEQ ID NO : 6 , or amino acid sequence of SEQ ID NO : 6 , or m . a Tryptophan to Cysteine amino acid substitution at an b . a Tryptophan to Serine amino acid substitution at an amino acid position 1999 (W1999C ) aligning with the amino acid position 1999 (W1999S ) aligning with the amino acid sequence of SEQ ID NO : 6 and an alanine amino acid sequence of SEQ ID NO : 6 , or to valine amino acid substitution at an amino acid c . an alanine to valine amino acid substitution at an amino position 2004 ( A2004V ) aligning with the amino acid acid position 2004 (A2004V ) aligning with the amino sequence of SEQ ID NO : 6 and a Tryptophan to Serine acid sequence of SEQ ID NO : 6 , or amino acid substitution at an amino acid position 2027 d . a Tryptophan to Serine amino acid substitution at an (W2027S ) aligning with the amino acid sequence of amino acid position 2027 (W2027S ) aligning with the SEQ ID NO : 6 , or amino acid sequence of SEQ ID NO : 6 , or n . a Tryptophan to Serine amino acid substitution at an e . a Tryptophan to Cysteine amino acid substitution at an amino acid position 1999 (W1999S ) aligning with the amino acid position 1999 (W1999C ) aligning with the amino acid sequence of SEQ ID NO : 6 and an alanine amino acid sequence of SEQ ID NO : 6 and a Trypto to valine amino acid substitution at an amino acid phan to Serine amino acid substitution at an amino acid position 2004 (A2004V ) aligning with the amino acid position 1999 (W1999S ) aligning with the amino acid sequence of SEQ ID NO : 6 and a Tryptophan to Serine sequence of SEQ ID NO : 6 , or amino acid substitution at an amino acid position 2027 f . a Tryptophan to Cysteine amino acid substitution at an (W2027S ) aligning with the amino acid sequence of amino acid position 1999 (W1999C ) aligning with the SEQ ID NO : 6 . amino acid sequence of SEQ ID NO : 6 and an alanine 4 . ( canceled ) to valine amino acid substitution at an amino acid 5 . ( canceled ) position 2004 (A2004V ) aligning with the amino acid 6 . The sorghum plant or plant part of claim 1 , wherein the sequence of SEQ ID NO : 6 , or plant or plant part is homozygous or heterozygous for one or g . a Tryptophan to Cysteine amino acid substitution at an more of mutation of the ACC gene . amino acid position 1999 (W1999C ) aligning with the 7 . The sorghum plant or plant part of claim 1 , wherein the amino acid sequence of SEQ ID NO : 6 and a Trypto plant or plant part comprises one or more mutations of the phan to Serine amino acid substitution at an amino acid ACC gene in homozygous or heterozygous combinations . position 2027 W2027S ) aligning with the amino acid 8 . The sorghum plant part of claim 1 , wherein the plant sequence of SEQ ID NO : 6 , or part is an organ , tissue , cell or seed . h . a Tryptophan to Serine amino acid substitution at an 9 . One or more Acetyl- CoA carboxylase (ACC ) inhibiting amino acid position 1999 (W1999S ) aligning with the herbicide capable of being used for controlling unwanted amino acid sequence of SEQ ID NO : 6 and an alanine vegetation in one or more sorghum growing areas , wherein to valine amino acid substitution at an amino acid position 2004 (A2004V ) aligning with the amino acid the sorghum plants in the growing area comprise one or sequence of SEQ ID NO : 6 , or more ACC inhibitor herbicide tolerant sorghum plants of i. a Tryptophan to Serine amino acid substitution at an claim 1 . amino acid position 1999 (W1999S ) aligning with the 10 . ( canceled ) amino acid sequence of SEQ ID NO : 6 and a Trypto 11 . A method for introducing creating an Acetyl- CoA phan to Serine amino acid substitution at an amino acid carboxylase (ACC ) inhibitor herbicide tolerant sorghum position 2027 W2027S ) aligning with the amino acid plant or plant part having one or more mutations in the sequence of SER ID NO : 6 , or Acetyl- CoA Carboxylase (ACC ) gene comprising the steps j . an alanine to valine amino acid substitution at an amino of: acid position 2004 ( A2004V ) aligning with the amino a . exposing a sorghum plant or plant part to about 1 acid sequence of SEQ ID NO : 6 and a Tryptophan to uM -200 UM of an ACC inhibitor herbicide , Serine amino acid substitution at an amino acid posi b . selecting a cell, plant or plant part which grows in the tion 2027 (W2027S ) aligning with the amino acid presence of up to 200 uM of an ACC inhibitor herbi sequence of SEQ ID NO : 6 , or cide , and k . a Tryptophan to Cysteine amino acid substitution at an c . regenerating plant shoots from the selected cell, plant or amino acid position 1999 (W1999C ) aligning with the plant part in the presence of an ACC inhibitor herbi amino acid sequence of SEQ ID NO : 6 and a Trypto cide . phan to Serine amino acid substitution at an amino acid 12 . (canceled ) position 1999 (W1999S ) aligning with the amino acid 13 . A method of creating an Acetyl- CoA carboxylase sequence of SEQ ID NO : 6 and an alanine to valine ( ACC ) herbicide tolerant sorghum plant or plant part having amino acid substitution at an amino acid position 2004 one or more mutations in the Acetyl - CoA Carboxylase (A2004V ) aligning with the amino acid sequence of ( ACC ) gene , comprising the steps of SEQ ID NO : 6 , or a . mutating the endogenous nucleotide sequence encoding 1. a Tryptophan to Cysteine amino acid substitution at an the ACC protein by inserting, deleting , modifying or amino acid position 1999 (W1999C ) aligning with the replacing one ormore nucleotides within the genome of amino acid sequence of SEQ ID NO : 6 and a Trypto living sorghum tissue using an engineered nuclease that US 2018 /0346920 A1 Dec . 6 , 2018 40

creates site - specific double - strand breaks (DSBs ) at a f. 50 % or greater desired location in the genome, g . 60 % or greater . b . selecting a cell, plant or plant part comprising the 21 . ( canceled ) mutation and wherein the plant or plant part grows in 22 . A method of producing ACC inhibitor herbicide the presence of up to 200 uM of an ACC inhibitor tolerant sorghum plant progeny comprising the steps of a . crossing a first ACC inhibitor herbicide tolerant sor herbicide , and ghum plant of claim 1 with a second sorghum plant c . regenerating plant shoots from the selected cell , plant or having a different genetic background , plant part in the presence of an ACC inhibitor herbi b . selecting a progeny plant resulting from the crossing cide . wherein the progeny comprises the mutation in the 14 . The method of claim 13 , wherein the endogenous ACC gene of the first ACC inhibitor herbicide tolerant nucleotide sequence encoding the ACC protein is mutating sorghum plant . using Meganuclease, Zinc - Fingure Nuclease , TALEN , or 23. - 25 . (canceled ) CRISPR / Cas9 technologies . 26 . A method of developing a population of Acetyl- CoA 15 . A method of creating an Acetyl - CoA carboxylase carboxylase (ACC ) inhibitor herbicide tolerant sorghum (ACC ) herbicide tolerant sorghum plant or plant part having plants comprising the steps of one or more mutations in the Acetyl- CoA Carboxylase a . screening a population of sorghum plants to identify a (ACC ) gene , comprising the steps of plant comprising the mutation of the an ACC inhibitor a . transforming a plant cell with one or more expression herbicide tolerant sorghum plant of claim 1 , and vectors , wherein the expression vector comprises a b . propagating the identified sorghum plants comprising a transgene nucleotide sequence , wherein the transgene mutation in the ACC gene nucleotide sequence to nucleotide sequence encodes a mutated ACC protein develop a population of ACC inhibitor herbicide toler amino acid sequence , ant sorghum plants . b . selecting a cell , plant or plant part that expresses the 27. - 36 . ( canceled ) mutated AAC protein and grows in the presence of up 37 . A method for controlling unwanted vegetation in a to 200 uM of an ACC inhibitor herbicide , and sorghum plant growing area comprising an ACC inhibitor c . regenerating plant shoots from the selected cell, plant or herbicide tolerant sorghum plant of claim 1 with one or more plant part in the presence of an ACC inhibitor herbi ACC inhibitor herbicide ( s ) , wherein the AAC inhibitor cide . herbicide is applied alone or in combination with one or 16 . Themethod of claim 15 wherein the transgene nucleo more non - ACC inhibitor herbicide. tide sequence is derived from any source . 38 . The method of claim 37, wherein the AAC inhibitor 17 . The method of claim 15 wherein the plant cell is herbicide and the non - ACC inhibitor herbicide are applied transformed through PEG mediated protoplast transforma jointly or simultaneously . tion , protoplast electroporation , biolistics , or agrobacterium 39 . The method of claim 37 , wherein the AAC inhibitor mediated transformation . herbicide and the non - ACC inhibitor herbicide are applied at 18 . (canceled ) different times 19 . The method of claim 15 , wherein the plants shoots are 40 . The method of claim 37 , wherein the AAC inhibitor regenerated at an efficiency of herbicide and the non -ACC inhibitor herbicide are applied a . 25 % or greater, sequentially , in pre -emergence applications followed by b . 30 % or greater , post - emergence applications, or in early post - emergence c . 35 % or greater , applications followed by medium or late post - emergence d . 40 % or greater, applications. e . 45 % or greater, or 41. - 55 . (canceled ) f. 50 % or greater. 56 . The sorghum plant or plant part of claim 1 , wherein 20 . The method of claim 15 wherein the efficiency of the plant or plant part corresponds to the deposit under regenerating a sorghum plant is ATCC Accession No. PTA - 125106 , PTA - 125107 or PTA a . 25 % or greater, 125108 . b . 30 % or greater, 57 . A plant progeny of the plant of claim 56 . c . 35 % or greater , 58 . A seed corresponding to the deposit under ATCC d . 40 % or greater, Accession No. PTA - 125106 , PTA - 125107 or PTA - 125108 . e . 45 % or greater, or