# US 8,465,928 B2 · Canola event PV-BNGT04(RT73) and compositions and methods for detection thereof

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- Number: US 8,465,928 B2
- Document ID: US8465928B2
- Status: Granted
- Kind: B2
- Application number: 13/081,494
- Filed: 2011-04-06
- Published: 2011-10-20
- Granted: 2013-06-18
- Earliest claimed: 2000-10-30
- Assignees: Monsanto Technology LLC
- Applicants: Rachel Krieb; Qingyi Zeng
- Inventors: Rachel Krieb; Qingyi Zeng
- Web page: https://patentgrep.com/patent/US8465928B2

## CPC

- C07H 21/04: with deoxyribosyl as saccharide radical
- C12N 15/8275: {Glyphosate}
- C12Q 1/6876: Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q 1/6895: for plants, fungi or algae
- Y10T 436/143333: Saccharide [e.g., DNA, etc.]
- C07H: SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS (derivatives of aldonic or saccharic acids C07C, C07D; aldonic acids, saccharic acids C07C59/105, C07C59/285; cyanohydrins C07C255/16; glycals C07D; compounds of unknown constitution C07G; polysaccharides, derivatives thereof C08B; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; sugar industry C13)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- 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
- Y10T: TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION

## Abstract

The present invention provides assays for detecting the presence of the PV-BNGT04(RT73) canola event based on the DNA sequence of the recombinant construct inserted into the canola genome and of genomic sequences flanking the insertion site.

## Claims

### Claim 1

A DNA molecule comprising a DNA sequence selected from the group consisting of SEQ ID NO: 7 and SEQ ID NO: 8.

### Claim 2

A method of detecting the presence of DNA corresponding to the genomic/transgene DNA of canola PV-BNGT04(RT73) event and progeny thereof in a sample, the method comprising:
(a) contacting the sample comprising canola DNA with a primer pair that when used in a nucleic acid amplification reaction with DNA from canola PV-BNGT04(RT73), produces an amplicon that is diagnostic for canola PV-BNGT04(RT73); and
(b) performing the nucleic acid amplification reaction, thereby producing the amplicon; and
(c) detecting the amplicon.

### Claim 3 (depends on 2)

An isolated DNA molecule comprising the amplicon produced by the method of claim 2.

### Claim 4

A DNA detection kit specific for genomic/transgene DNA of canola PV-BNGT04(RT73) and progeny thereof comprising:
(a) a pair of DNA molecules, wherein a first molecule in said pair comprises 11 nucleotides or more of SEQ ID NO: 7, or a sequence fully complementary thereto, wherein said pair is capable of producing an amplicon diagnostic for event PV-BNGT04(RT73), wherein said amplicon comprises SEQ ID NO: 5; or
(b) a pair of DNA molecules, wherein a first molecule in said pair comprises 11 nucleotides or more of SEQ ID NO: 8, or a sequence fully complementary thereto, wherein said pair is capable of producing an amplicon diagnostic for event PV-BNGT04(RT73), wherein said amplicon comprises SEQ ID NO: 6.

### Claim 5

A method of detecting the presence of a genomic/transgene DNA corresponding to the PV-BNGT04(RT73) and progeny thereof in a sample, the method comprising:
(a) contacting the sample comprising canola DNA with a polynucleotide probe that hybridizes under stringent hybridization conditions with DNA from canola event PV-BNGT04(RT73) and progeny thereof and does not hybridize under the stringent hybridization conditions with a non-PV-BNGT04(RT73) and progeny thereof canola plant DNA; and
(b) subjecting the sample and probe to stringent hybridization conditions;
(c) detecting hybridization of the probe to the DNA.

### Claim 6

An isolated DNA molecule comprising a genomic/transgene DNA junction sequence of canola PV-BNGT04 or a sequence fully complementary thereto.

### Claim 7

A method of breeding a glyphosate tolerant trait into canola plants comprising:
(a) extracting a DNA sample from progeny canola plants;
(b) detecting in the DNA sample a nucleic acid molecule selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 6, and a sequence fully complementary thereto; and
(c) performing a marker assisted breeding method for the glyphosate tolerant trait, wherein the glyphosate tolerant trait is genetically linked to a complement of the marker nucleic acid molecule.