# US 10,480,006 B2 · Pesticidal genes and methods of use

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- Number: US 10,480,006 B2
- Document ID: US10480006B2
- Status: Granted
- Kind: B2
- Application number: 14/972,757
- Filed: 2015-12-17
- Published: 2016-06-23
- Granted: 2019-11-19
- Earliest claimed: 2014-12-22
- Assignees: AgBiome, Inc.
- Applicants: AgBiome, Inc.
- Inventors: Jessica Parks; Kira Bulazel Roberts; Rebecca E. Thayer
- Web page: https://patentgrep.com/patent/US10480006B2

## CPC

- C12N 15/8286: {for insect resistance}
- A01N 63/50: Isolated enzymes; Isolated proteins (peptides A01N37/46)
- C07K 14/325: Bacillus thuringiensis crystal peptides, i.e. delta-endotoxins
- C12N 15/8261: {with agronomic (input) traits, e.g. crop yield}
- Y02A 40/146: Genetically Modified [GMO] plants, e.g. transgenic plants
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- 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
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- Y02A: TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE

## Abstract

Compositions having pesticidal activity and methods for their use are provided. Compositions include isolated and recombinant polypeptides having pesticidal activity, recombinant and synthetic nucleic acid molecules encoding the polypeptides, DNA constructs and vectors comprising the nucleic acid molecules, host cells comprising the vectors, and antibodies to the polypeptides. Polynucleotide sequences encoding the polypeptides can be used in DNA constructs or expression cassettes for transformation and expression in organisms of interest. The compositions and methods provided are useful for producing organisms with enhanced pest resistance or tolerance. Transgenic plants and seeds comprising a nucleotide sequence that encodes a pesticidal protein of the invention are also provided. Such plants are resistant to insects and other pests. Methods are provided for producing the various polypeptides disclosed herein, and for using those polypeptides for controlling or killing a pest. Methods and kits for detecting polypeptides of the invention in a sample are also included.

## Claims

### Claim 1

A recombinant polypeptide, comprising
a polypeptide comprising an amino acid sequence having at least 95% sequence identity to the amino acid sequence as set forth in SEQ ID NO: 36, wherein said polypeptide has pesticidal activity and further comprises a heterologous amino acid sequence chemically linked to said polypeptide.

### Claim 2 (depends on 1)

A composition comprising the polypeptide of claim 1.

### Claim 3

A recombinant nucleic acid molecule encoding an amino acid sequence comprising
at least 95% sequence identity to the amino acid sequence as set forth in SEQ ID NO:36.

### Claim 4 (depends on 3)

The recombinant nucleic acid of claim 3, wherein said nucleic acid molecule is a synthetic sequence designed for expression in a plant.

### Claim 5 (depends on 3)

The recombinant nucleic acid molecule of claim 3, wherein said heterologous promoter is capable of directing expression in a plant cell.

### Claim 6 (depends on 3)

The recombinant nucleic acid molecule of claim 3, wherein said heterologous promoter is capable of directing expression in a bacteria.

### Claim 7 (depends on 3)

A host cell comprising the recombinant nucleic acid molecule of claim 3.

### Claim 8 (depends on 7)

The host cell of claim 7, wherein said host cell is a bacterial host cell.

### Claim 9

A DNA construct comprising a heterologous promoter that drives expression in a plant cell operably linked to a nucleotide sequence that encodes
a polypeptide comprising the amino acid sequence having at least 95% sequence identity to an amino acid sequence as set forth in SEQ ID NO:36.

### Claim 10 (depends on 9)

The DNA construct of claim 9, wherein said nucleotide sequence is a synthetic DNA sequence designed for expression in a plant.

### Claim 11 (depends on 9)

A vector comprising the DNA construct of claim 9.

### Claim 12 (depends on 11)

A host cell comprising the DNA construct of claim 11.

### Claim 13 (depends on 12)

A composition comprising the host cell of claim 12.

### Claim 14 (depends on 13)

The composition of claim 13, wherein said composition is selected from the group consisting of a powder, dust, pellet, granule, spray, emulsion, colloid, and solution.

### Claim 15 (depends on 14)

The composition of claim 14, wherein said composition comprises from about 1% to about 99% by weight of said polypeptide.

### Claim 16 (depends on 2)

A method for controlling a pest population comprising contacting said pest population with a pesticidal-effective amount of the composition of claim 2.

### Claim 17 (depends on 12)

A method for producing a polypeptide with pesticidal activity comprising culturing the host cell of claim 12 under conditions in which the nucleic acid molecule encoding the polypeptide is expressed.

### Claim 18

A plant having stably incorporated into its genome a DNA construct comprising
a nucleotide sequence that encodes a polypeptide comprising an amino acid sequence having at least 95% sequence identity to the amino acid sequence as set forth in SEQ ID NO:36, wherein said polypeptide has pesticidal activity.

### Claim 19 (depends on 18)

A transgenic seed of the plant of claim 18.

### Claim 20

A method for protecting a plant from an insect pest, comprising expressing in a plant or cell thereof
a nucleotide sequence that encodes a polypeptide comprising an amino acid sequence having at least 95% sequence identity to the amino acid sequence as set forth in SEQ ID NO:36, wherein said nucleotide sequence encodes a polypeptide having pesticidal activity and is operably linked to a promoter capable of driving expression in the plant or cell.

### Claim 21 (depends on 20)

The method of claim 20, wherein said plant produces a pesticidal polypeptide having pesticidal activity against a Hemipteran pest.

### Claim 22 (depends on 1)

The recombinant polypeptide of claim 1, wherein the polypeptide comprises an amino acid sequence having at least 90% sequence identity to the amino acid sequence set forth in SEQ ID NO: 35.

### Claim 23 (depends on 22)

A composition comprising the polypeptide of claim 22.

### Claim 24 (depends on 3)

The recombinant nucleic acid molecule of claim 3, wherein the recombinant nucleic acid molecule encodes an amino acid sequence having at least 90% sequence identity to the amino acid sequence set forth in SEQ ID NO: 35.

### Claim 25 (depends on 24)

A host cell comprising the recombinant nucleic acid molecule of claim 24.

### Claim 26 (depends on 9)

The DNA construct of claim 9, wherein the nucleotide sequence encodes an amino acid sequence having at least 90% sequence identity to the amino acid sequence set forth in SEQ ID NO: 35.

### Claim 27 (depends on 26)

A vector comprising the DNA construct of claim 26.

### Claim 28 (depends on 26)

A host cell comprising the DNA construct of claim 26.

### Claim 29 (depends on 28)

A composition comprising the host cell of claim 28.

### Claim 30 (depends on 18)

The plant of claim 18, wherein the nucleotide sequence encodes a polypeptide comprising an amino acid sequence having at least 90% sequence identity to the amino acid sequence set forth in SEQ ID NO: 35.

### Claim 31 (depends on 30)

A transgenic seed of the plant of claim 30.

### Claim 32 (depends on 23)

A method for controlling a pest population comprising contacting said pest population with a pesticidal-effective amount of the composition of claim 23.

### Claim 33 (depends on 25)

A method for producing a polypeptide with pesticidal activity comprising culturing the host cell of claim 25 under conditions in which the nucleic acid molecule encoding the polypeptide is expressed.

### Claim 34 (depends on 20)

The method of claim 20, wherein the nucleotide sequence encodes a polypeptide comprises an amino acid sequence having at least 90% sequence identity to the amino acid sequence set forth in SEQ ID NO: 35.

### Claim 35 (depends on 1)

The recombinant polypeptide of claim 1, wherein the polypeptide comprises the amino acid sequence set forth in SEQ ID NO:36.

### Claim 36 (depends on 3)

The recombinant nucleic acid molecule of claim 3, wherein the nucleic acid molecule encodes the amino acid sequence set forth in SEQ ID NO:36.

### Claim 37 (depends on 9)

The DNA construct of claim 9, wherein the nucleotide sequence encodes a polypeptide comprising the amino acid sequence set forth in SEQ ID NO:36.

### Claim 38 (depends on 18)

The plant of claim 18, wherein the DNA construct comprises a nucleotide sequence that encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:36.

### Claim 39 (depends on 20)

The method of claim 20, wherein said nucleotide sequence encodes a polypeptide comprising the amino acid sequence of SEQ ID NO:36.