Herbicide tolerant soybean plants and methods for identifying same

Patent metadata

Field	Value
Publication number	US-9683242-B2
Application number	US-201013511631-A
Country	US
Kind code	B2
Filing date	Nov 23, 2010
Priority date	Nov 23, 2009
Publication date	Jun 20, 2017
Grant date	Jun 20, 2017

Abstract

The invention provides specific transgenic soybean plants, plant material and seeds, characterized in that these products harbor a stack of specific transformation events at specific locations in the soybean genome. Tools are also provided which allow rapid and unequivocal identification of these events in biological samples.

First claim

The invention claimed is:

1. A method for identifying the simultaneous presence of elite events EE-GM3 and EE-GM2 in biological samples, confirming seed purity in seed samples, or screening seeds for the presence of elite events EE-GM3 and EE-GM2 in samples in seed lots, which method comprises detection of an EE-GM3 specific region with a specific primer pair or probe which specifically recognizes the 5′ or 3′ flanking region of foreign DNA comprising herbicide tolerance genes in EE-GM3 and the inserted foreign DNA contiguous therewith, and detecting an EE-GM2 specific region with a specific primer pair or probe which specifically recognizes the 5′ or 3′ flanking region of foreign DNA comprising herbicide tolerance genes in EE-GM2 and the inserted foreign DNA contiguous therewith.

2. The method of claim 1 , said method comprising amplifying two DNA fragments of between 50 and 1000 bp from a nucleic acid present in said biological samples using a first polymerase chain reaction with at least two primers, one of said primers recognizing the 5′ flanking region of foreign DNA comprising herbicide tolerance genes in EE-GM3, said 5′ flanking region comprising the nucleotide sequence of SEQ ID No. 2 from nucleotide 1 to nucleotide 1451 or the 3′ flanking region of foreign DNA comprising herbicide tolerance genes in EE-GM3, said 3′ flanking region comprising the nucleotide sequence of the complement of SEQ ID No. 3 from nucleotide 241 to nucleotide 1408, the other primer of said primers recognizing a sequence within the foreign DNA of EE-GM3 comprising the nucleotide sequence of the complement of SEQ ID No. 2 from nucleotide 1452 to nucleotide 1843 or the nucleotide sequence of SEQ ID No. 3 from nucleotide 1 to nucleotide 240 or the other primer of said primers recognizing a sequence within the foreign DNA of EE-GM3 comprising the nucleotide sequence of SEQ ID No. 1 or its complement, or comprising the nucleotide sequence of SEQ ID No. 20 from nucleotide position 1452 to nucleotide position 16638 or its complement, and using a second polymerase chain reaction with at least two primers, one of said primers recognizing the 5′ flanking region of foreign DNA comprising herbicide tolerance genes in EE-GM2, said 5′ flanking region comprising the nucleotide sequence of SEQ ID No. 14 from nucleotide 1 to nucleotide 311 or the 3′ flanking region of foreign DNA comprising herbicide tolerance genes in EE-GM2, said 3′ flanking region comprising the nucleotide sequence of the complement of SEQ ID No. 15 from nucleotide 508 to nucleotide 1880, the other primer of said primers recognizing a sequence within the foreign DNA of EE-GM2 comprising the nucleotide sequence of the complement of SEQ ID No. 14 from nucleotide 312 to nucleotide 810 or the nucleotide sequence of SEQ ID No. 15 from nucleotide 1 to nucleotide 507 or the other primer of said primers recognizing a sequence within the foreign DNA of EE-GM2 comprising the nucleotide sequence of SEQ ID No. 11 or its complement, whereby said first and second polymerase reaction may be sequential or simultaneous.

3. The method of claim 2 ,
(a) wherein said primer recognizing the 5′ flanking region of EE-GM3 comprises a nucleotide sequence of 17 to 200 consecutive nucleotides selected from the nucleotide sequence of SEQ ID No. 2 from nucleotide 1 to nucleotide 1451 or said primer recognizing the 3′ flanking region of EE-GM3 comprises a nucleotide sequence of 17 to 200 consecutive nucleotides selected from the nucleotide sequence of the complement of SEQ ID No. 3 from nucleotide 241 to nucleotide 1408, and said primer recognizing a sequence within the foreign DNA of EE-GM3 comprises 17 to 200 consecutive nucleotides selected from the nucleotide sequence of the complement of SEQ ID No. 2 from nucleotide 1452 to nucleotide 1843 or the nucleotide sequence of SEQ ID No. 3 from nucleotide 1 to nucleotide 240 or the nucleotide sequence of SEQ ID No. 1 or its complement, or the nucleotide sequence of SEQ ID No. 20 from nucleotide position 1452 to nucleotide position 16638 or its complement, and wherein said primer recognizing the 5′ flanking region of EE-GM2 comprises a nucleotide sequence of 17 to 200 consecutive nucleotides selected from the nucleotide sequence of SEQ ID No. 14 from nucleotide 1 to nucleotide 311 or said primer recognizing the 3′ flanking region of EE-GM3 comprises a nucleotide sequence of 17 to 200 consecutive nucleotides selected from the nucleotide sequence of the complement of SEQ ID No. 15 from nucleotide 508 to nucleotide 1880, and said primer recognizing a sequence within the foreign DNA of EE-GM2 comprises 17 to 200 consecutive nucleotides selected from the nucleotide sequence of the complement of SEQ ID No. 14 from nucleotide 312 to nucleotide 810 or the nucleotide sequence of SEQ ID No. 15 from nucleotide 1 to nucleotide 507 or the nucleotide sequence of SEQ ID No. 11 or its complement; or
(b) wherein said primer recognizing the 5′ flanking region of EE-GM3 comprises at its extreme 3′ end a nucleotide sequence of at least 17 consecutive nucleotides selected from the nucleotide sequence of SEQ ID No. 2 from nucleotide 1 to nucleotide 1451 or said primer recognizing the 3′ flanking region of EE-GM3 comprises at its extreme 3′ end a nucleotide sequence of at least 17 consecutive nucleotides selected from the nucleotide sequence of the complement of SEQ ID No. 3 from nucleotide 241 to nucleotide 1408, and said primer recognizing a sequence within the foreign DNA of EE-GM3 comprises at its 3′ end at least 17 consecutive nucleotides selected from the nucleotide sequence of the complement of SEQ ID No. 2 from nucleotide 1452 to nucleotide 1843 or the nucleotide sequence of SEQ ID No. 3 from nucleotide 1 to nucleotide 240 or the nucleotide sequence of SEQ ID No. 1 or its complement, or the nucleotide sequence of SEQ ID No. 20 from nucleotide position 1452 to nucleotide position 16638 or its complement, and wherein said primer recognizing the 5′ flanking region of EE-GM2 comprises at its extreme 3′ end a nucleotide sequence of at least 17 consecutive nucleotides selected from the nucleotide sequence of SEQ ID No. 14 from nucleotide 1 to nucleotide 311 or said primer recognizing the 3′ flanking region of EE-GM2 comprises at its extreme 3′ end a nucleotide sequence of at least 17 consecutive nucleotides selected from the nucleotide sequence of the complement of SEQ ID No. 15 from nucleotide 508 to nucleotide 1880, and said primer recognizing a sequence within the foreign DNA of EE-GM2 comprises at its 3′ end at least 17 consecutive nucleotides selected from the nucleotide sequence of the complement of SEQ ID No. 14 from nucleotide 312 to nucleotide 810 or the nucleotide sequence of SEQ ID No. 15 from nucleotide 1 to nucleotide 507 or the nucleotide sequence of SEQ ID No. 11 or its complement.

4. The method of claim 3 , wherein said EE-GM3 specific primers comprise the sequence of SEQ ID No. 5 and SEQ ID No. 4, respectively, or the sequence of SEQ ID No. 7 and SEQ ID No. 5 respectively, and said EE-GM2 specific primers comprise the sequence of SEQ ID No. 18 and SEQ ID No. 19 respectively.

5. The method of claim 4 , which method comprises amplifying an EE-GM3 specific fragment of about 263 or 706 bp using the EE-GM3 PCR Identification Protocol and an EE-GM2 specific fragment of about 151 bp.

6. A kit for identifying the simultaneous presence of elite event EE-GM3 and elite event EE-GM2 in biological samples, said kit comprising one primer recognizing the 5′ flanking region of foreign DNA comprising herbicide tolerance genes in EE-GM3, said 5′ flanking region comprising the nucleotide sequence of SEQ ID No. 2 from nucleotide 1 to nucleotide 1451 or one primer recognizing the 3′ flanking region of foreign DNA comprising herbicide tolerance genes in EE-GM3, said 3′ flanking region comprising the nucl

Assignees

• Mason Justin Thomas
• Lettow Leslie James
• Eby Mark Alan
• Eby William H
• Welz Gunter
• Verhaeghe Steven
• De Beuckeleer Marc
• Habex Veerle
• Ferrulo Jean-Marc
• Ms Technologies Llc
• Bayer Cropscience Nv

Inventors

• Mason Justin Thomas
• Lettow Leslie James
• Eby Mark Alan
• Eby William H
• Welz Gunter
• Verhaeghe Steven
• De Beuckeleer Marc
• Habex Veerle
• Ferrulo Jean-Marc

Classifications

• C07H21/04

  with deoxyribosyl as saccharide radical · CPC title

• C12N15/8274 Primary

  for herbicide resistance · CPC title

• C12N15/8275 Primary

  Glyphosate · CPC title

• A01H5/10

  Seeds · CPC title

• C12Q1/6895 Primary

  for plants, fungi or algae · CPC title