# US 12,180,489 B2 · Modified excisable MON87708 soybean transgenic herbicide resistance locus

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- Number: US 12,180,489 B2
- Document ID: US12180489B2
- Status: Granted
- Kind: B2
- Application number: 18/058,144
- Filed: 2022-11-22
- Published: 2023-03-16
- Granted: 2024-12-31
- Earliest claimed: 2020-07-31
- Assignees: Inari Agriculture Technology, Inc.
- Applicants: Inari Agriculture Technology, Inc.
- Inventors: Michael Lee Nuccio; Michael Andreas Kock; Joshua L. Price
- Web page: https://patentgrep.com/patent/US12180489B2

## CPC

- C12N 9/22: Ribonucleases {[RNase]; Deoxyribonucleases [DNase]}
- C12N 15/8201: {Methods for introducing genetic material into plant cells, e.g. DNA, RNA, stable or transient incorporation, tissue culture methods adapted for transformation}
- A01H 1/02: Methods or apparatus for hybridisation; Artificial pollination {; Fertility}
- A01H 5/10: Seeds
- A01H 6/4684: {Zea mays [maize]}
- A01H 6/542: {Glycine max [soybean]}
- C07K 14/415: from plants
- C12N 9/226: {Class 2 CAS enzyme complex, e.g. single CAS protein}
- C12N 15/11: DNA or RNA fragments; Modified forms thereof (DNA or RNA not used in recombinant technology, C07H21/00); {Non-coding nucleic acids having a biological activity}
- C12N 15/8213: {Targeted insertion of genes into the plant genome by homologous recombination}
- C12N 15/8286: {for insect resistance}
- C12Q 1/6834: Enzymatic or biochemical coupling of nucleic acids to a solid phase
- C12Q 1/6895: for plants, fungi or algae
- C12N 2310/20: involving clustered regularly interspaced short palindromic repeats [CRISPR]
- C12N 2800/80: Vectors containing sites for inducing double-stranded breaks, e.g. meganuclease restriction sites
- C12Q 2600/13: Plant traits
- C12Q 2600/156: Polymorphic or mutational markers
- C12Q 2600/158: Expression markers
- 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)
- A01H: NEW PLANTS OR {NON-TRANSGENIC} PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- 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)
- 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
- Y02A: TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE

## Abstract

Transgenic INHT30 soybean plants comprising modifications of the MON87708 soybean locus which provide for facile excision of the modified MON87708 transgenic locus or portions thereof, methods of making such plants, and use of such plants to facilitate breeding are disclosed.

## Claims

### Claim 1

A transgenic soybean plant cell comprising an INHT30 transgenic locus comprising the DNA molecule set forth in SEQ ID NO: 23.

### Claim 2 (depends on 1)

A transgenic soybean plant part comprising the soybean plant cell of claim 1.

### Claim 3 (depends on 2)

The transgenic soybean plant part of claim 2, wherein said soybean plant part is a seed.

### Claim 4 (depends on 1)

A transgenic soybean plant comprising the soybean plant cell of claim 1.

### Claim 5 (depends on 4)

A method for obtaining a bulked population of inbred seed comprising selfing the transgenic soybean plant of claim 4 and harvesting seed comprising the INHT30 transgenic locus from the selfed soybean plant.

### Claim 6 (depends on 4)

A method of obtaining hybrid soybean seed comprising crossing the transgenic soybean plant of claim 4 to a second soybean plant which is genetically distinct from the first soybean plant and harvesting seed comprising the INHT30 transgenic locus from the cross.

### Claim 7

A DNA molecule comprising SEQ ID NO: 23.

### Claim 8 (depends on 7)

A processed transgenic soybean plant product comprising the DNA molecule of claim 7.

### Claim 9 (depends on 7)

A biological sample containing the DNA molecule of claim 7.

### Claim 10 (depends on 1)

A method of detecting a soybean plant cell comprising the INHT30 transgenic locus of claim 1, comprising the step of detecting DNA molecule comprising SEQ ID NO: 8.

### Claim 11 (depends on 1)

A method of excising the INHT30 transgenic locus from the genome of the soybean plant cell of claim 1, comprising the steps of: (a) contacting the INHT30 transgenic locus of the plant cell with: (i) a Cas12a RNA dependent DNA endonuclease (RdDe); and (ii) a guide RNA (gRNA) capable of hybridizing to the guide RNA hybridization site of the originator guide RNA recognition site (OgRRS) and the cognate guide RNA recognition site (CgRRS) of SEQ ID NO: 23; wherein the Cas12a RdDe recognizes a OgRRS/gRNA and a CgRRS/gRNA hybridization complex; and, (b) selecting a transgenic plant cell, transgenic plant part, or transgenic plant wherein the INHT30 transgenic locus flanked by the OgRRS and the CgRRS has been excised.

### Claim 12 (depends on 11)

The method of claim 11, wherein said guide RNA comprises an RNA sequence encoded by SEQ ID NO: 13.