# US 10,370,678 B2 · Acetyl co-enzyme A carboxylase herbicide resistant plants

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- Number: US 10,370,678 B2
- Document ID: US10370678B2
- Status: Granted
- Kind: B2
- Application number: 15/401,500
- Filed: 2017-01-09
- Published: 2017-06-15
- Granted: 2019-08-06
- Earliest claimed: 2011-02-01
- Assignees: Colorado Wheat Research Foundation, Inc.
- Applicants: Colorado Wheat Research Foundation, Inc.
- Inventors: Michael Hal Ostlie; Scott Haley; Philip Westra; Victoria Ashley Anderson
- Web page: https://patentgrep.com/patent/US10370678B2

## CPC

- C12N 15/8274: {for herbicide resistance}
- A01H 5/10: Seeds
- A01H 6/4678: {Triticum sp. [wheat]}
- A01N 43/54: 1,3-Diazines; Hydrogenated 1,3-diazines
- C12N 9/93: {Ligases (6)}
- C12N 15/8218: {Antisense, co-suppression, viral induced gene silencing [VIGS], post-transcriptional induced gene silencing [PTGS]}
- C12Q 1/6895: for plants, fungi or algae
- C12Y 604/01002: Acetyl-CoA carboxylase (6.4.1.2)
- C12Q 2600/13: Plant traits
- C12Q 2600/156: Polymorphic or mutational markers
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- A01H: NEW PLANTS OR {NON-TRANSGENIC} PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01N: PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF (preservation of food or foodstuff A23); BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES (preparations for medical, dental or toiletry purposes which kill or prevent the growth or proliferation of unwanted organisms A61K); PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Y: ENZYMES

## Abstract

The present invention provides for compositions and methods for producing crop plants that are resistant to herbicides. In particular, the present invention provides for wheat plants, plant tissues and plant seeds that contain altered acetyl-CoA carboxylase (ACCase) genes and proteins that are resistant to inhibition by herbicides that normally inhibit the activity of the ACCase protein.

## Claims

### Claim 1

A monocot cereal plant having an acetyl-CoA carboxylase protein which confers resistance to one or more acetyl-CoA carboxylase herbicides at levels of said one or more herbicides that would inhibit the growth of said plant, wherein said acetyl-CoA carboxylase protein comprises a mutation at a position corresponding to position 2004 of the black grass reference sequence, SEQ ID NOS: 14 or 16.

### Claim 2 (depends on 1)

The monocot cereal plant of claim 1, wherein said one or more acetyl-CoA carboxylase herbicides are from a group consisting of aryloxyphenoxypropionates, cyclohexanediones, and phenylpyrazolin (DENs).

### Claim 3 (depends on 1)

The monocot cereal plant of claim 1, wherein said resistance to inhibition by one or more acetyl-CoA carboxylase herbicides is introduced into said plant by breeding.

### Claim 4 (depends on 1)

The monocot cereal plant of claim 1, wherein said acetyl-CoA carboxylase protein confers resistance to inhibition by one or more acetyl CoA carboxylase herbicides comprises an alanine-to-valine substitution Ala2004Val at a position corresponding to position 2004 of the black grass reference sequence, SEQ ID NOS: 14 or 16.

### Claim 5 (depends on 4)

The monocot cereal plant of claim 4, wherein said protein is SEQ ID NO: 8, 10, or 12 or its conservatively modified variants that include the ALA2004Val substitution.

### Claim 6 (depends on 1)

The monocot cereal plant of claim 1, wherein said plant is selected form wheat, barely, triticale, rice or maize.

### Claim 7 (depends on 6)

The monocot cereal plant of claim 6, wherein said plant is wheat.

### Claim 8 (depends on 7)

The wheat plant of claim 7, wherein said wheat plant comprises the nucleic acid sequence of SEQ ID NO: 4, 5, or 6.

### Claim 9 (depends on 8)

The wheat plant of claim 8, wherein said SEQ ID NO: 4, 5, or 6 is introduced into said wheat plant germplasm by breeding.

### Claim 10 (depends on 1)

The monocot cereal plant of claim 1, wherein said plant is created by introducing to said plant a heterologous nucleic acid sequence which encodes an ACCase protein with one or more substitutions that confer resistance to one or more acetyl-CoA carboxylase herbicides, wherein at least one of the substitutions is at a position corresponding to position 2004 when referenced to of the black grass reference sequence, SEQ ID NOS: 14 or 16.

### Claim 11 (depends on 1)

A method of controlling weeds in the vicinity of a monocot cereal plant of claim 1, comprising:
a. providing one or more acetyl-CoA carboxylase herbicides,
b. applying said one or more acetyl-CoA carboxylase herbicides to a field comprising the monocot cereal plant of claim 1, wherein weed growth is adversely affected by the application of said one or more herbicides and growth of said monocot cereal plant is not adversely affected.

### Claim 12 (depends on 11)

The method of claim 11, wherein said one or more acetyl-CoA carboxylase herbicides are from a group consisting of aryloxyphenoxypropionates, cyclohexanediones, and phenylpyrazolin (DENs).

### Claim 13 (depends on 1, 11)

The method of claim 11, wherein said monocot cereal plant of claim 1 comprises the resistance to inhibition by one or more acetyl-CoA carboxylase herbicides is introduced into the monocot cereal plant by introgression.

### Claim 14 (depends on 11)

The method of claim 11, wherein said mutation in the acetyl-CoA carboxylase protein is an amino acid substitution Ala2004Val in reference to SEQ ID NO: 14 or 16.

### Claim 15 (depends on 11)

The method of claim 11, wherein said monocot cereal plant comprises SEQ ID NO: 4.

### Claim 16 (depends on 11)

The method of claim 11, wherein said monocot cereal plant comprises one or more mutations in the acetyl-CoA carboxylase gene as found in AF28-A, AF26-B and/or AF10-D, wherein representative samples of seeds of AF28-A, AF26-B and AF10-D have been deposited under ATCC Nos PTA-123074, PTA-123076, and PTA-123075, respectively.

### Claim 17 (depends on 16)

The method of claim 16, wherein said acetyl-CoA carboxylase gene as found in AF28-A, AF26-B and AF10-D is introduced into said monocot cereal plant by breeding, wherein representative samples of seeds of AF28-A, AF26-B and AF10-D have been deposited under ATCC Nos PTA-123074, PTA-123076, and PTA-123075, respectively.

### Claim 18 (depends on 11)

The method of claim 11, wherein said monocot cereal plant is created by introducing to said plant a heterologous nucleic acid sequence which encodes an ACCase protein with one or more substitutions which confers resistance to one or more acetyl-CoA carboxylase herbicides, wherein one of the substitutions is at the position corresponding to position 2004 of the black grass reference sequence, SEQ ID NOS: 14 or 16.

### Claim 19

A method for producing a plant resistant to one or more acetyl-CoA carboxylase herbicides comprising:
a) introducing to said plant a nucleic acid sequence which encodes an ACCase protein that comprises an amino acid substitution at a position corresponding to position 2004 of the black grass reference sequence SEQ ID NO: 14 or 16 and wherein said ACCase protein confers resistance to inhibition by one or more acetyl-CoA carboxylase herbicides at levels of said one or more herbicides that would inhibit the growth of a plant.

### Claim 20 (depends on 19)

The method of claim 19, wherein said introducing is by backcrossing.

### Claim 21 (depends on 19)

The method of claim 19, wherein said introducing to said plant is transformation with a heterologous nucleic acid sequence encoding the ACCase protein that comprises an alanine-to-valine substitution Ala2004Val at the position 2004 of the black grass reference sequence SEQ ID NO: 14 or 16.

### Claim 22 (depends on 20)

The method of claim 20, wherein said introducing comprises b) obtaining a plant AF28-A, AF26-B and/or AF10-D, crossing said plant with an elite plant to create progeny plants, c) selecting plants with ACCase herbicide resistance, and repeating steps b and c so that the morphological and physiological characteristics of said elite plant are regenerated in a progeny plant that also has ACCase herbicide resistance, wherein representative samples of seeds of AF28-A, AF26-B and AF10-D have been deposited under ATCC Nos PTA-123074, PTA-123076, and PTA-123075, respectively.

### Claim 23 (depends on 19)

The method of claim 19, wherein said herbicide resistant plant comprises mutations in the acetyl-CoA carboxylase gene as found in AF28-A, AF26-B and/or AF10-D, wherein representative samples of seeds of AF28-A, AF26-B and AF10-D have been deposited under ATCC Nos PTA-123074, PTA-123076, and PTA-123075, respectively.

### Claim 24

A plant with a modified ACCase polynucleotide sequence, said sequence encoding an ACCase polypeptide having a valine at the position corresponding to position 2004 of the black grass reference sequence SEQ ID NO: 14, or 16.

### Claim 25 (depends on 24)

A seed of the plant of claim 24, wherein the seed comprises the modified ACCase polynucleotide sequence.

### Claim 26 (depends on 25)

A plant, or a part thereof, produced by growing the seed of claim 25, wherein the plant, or the part thereof, comprises the modified ACCase polynucleotide sequence.

### Claim 27 (depends on 26)

A tissue culture produced from protoplasts or cells from the plant of claim 26, wherein said cells or protoplasts are produced from a plant part selected from the group consisting of leaf, pollen, ovule, embryo, cotyledon, hypocotyl, meristematic cell, root, root tip, pistil, anther, flower, seed, shoot, stem, pod and petiole, wherein tissue culture comprises the modified ACCase polynucleotide sequence.

### Claim 28 (depends on 27)

A plant regenerated from the tissue culture of claim 27, wherein the regenerated plant comprises the modified ACCase polynucleotide sequence.

### Claim 29 (depends on 26)

A method for producing a seed, said method comprising crossing two plants and harvesting the resultant seed, wherein at least one plant is the plant of claim 26.

### Claim 30 (depends on 29)

A seed produced by the method of claim 29, wherein the seed comprises the modified ACCase polynucleotide sequence.

### Claim 31 (depends on 30)

A plant, or a part thereof, produced by growing said seed of claim 30, wherein the plant, or the part thereof, comprises the modified ACCase polynucleotide sequence.

### Claim 32 (depends on 29)

The method of claim 29, wherein at least one of said plants is a transgenic plant comprising the modified ACCase polynucleotide sequence.

### Claim 33

A recombinant polynucleotide comprising a nucleic acid sequence selected from the group consisting of:
(a) a nucleic acid sequence set forth in SEQ ID NO: 5, or 6;
(b) a nucleic acid nucleotide sequence that encodes a polypeptide having the amino acid sequence set forth in SEQ ID NO: 10, 11 or 12;
(c) a nucleic acid sequence having at least 95% sequence identity over the entire length of SEQ ID NO: 4, 5, or 6; and
(e) a nucleic acid sequence that comprises the full length complement of any one of (a) to (d);
wherein said nucleic acid sequence encodes a polypeptide with ACCase activity and resistance to an ACCase herbicide, wherein said polypeptide has at least 95% sequence identity to SEQ ID NO: 10, 11 or 12 and having a valine at a position corresponding to position 2004 of the black grass reference ACCase sequence SEQ ID NO:14, or 16; and
wherein said nucleic acid sequence is operably linked to a heterologous element.

### Claim 34 (depends on 33)

The recombinant polynucleotide of claim 33, wherein said polynucleotide is optimized for expression in a plant.

### Claim 35 (depends on 33)

A nucleic acid construct comprising the recombinant polynucleotide of claim 33, wherein said polynucleotide is operably linked to a promoter that drives expression in a host cell.

### Claim 36 (depends on 35)

An expression cassette comprising the nucleic acid construct of claim 35.

### Claim 37 (depends on 35)

A host cell having stably incorporated into its genome at least one nucleic acid construct of claim 35.

### Claim 38 (depends on 37)

The host cell of claim 37, wherein said host cell is a plant cell.

### Claim 39 (depends on 35)

A transgenic plant having stably incorporated into its genome the nucleic acid construct of claim 35.

### Claim 40 (depends on 39)

The transgenic plant according to claim 39, wherein said plant is selected from the group consisting of: corn, soybean, wheat, rice, alfalfa, barley, millet, sunflower, sorghum, canola and cotton.

### Claim 41 (depends on 40)

The transgenic plant according to claim 40, wherein said plant is wheat.