FAD2 performance loci and corresponding target site specific binding proteins capable of inducing targeted breaks

• US 9,963,711 B2
• Filed: 09/05/2013
• Issued: 05/08/2018
• Est. Priority Date: 09/07/2012
• Status: Active Grant

• Current Assignee: Dow Agrosciences LLC (Corteva, Inc.), Sangamo Therapeutics, Inc.
• Original Assignee: Dow Agrosciences LLC (Corteva, Inc.), Sangamo Therapeutics, Inc.
• Inventors: Cogan, Noel, Forster, John, Hayden, Matthew, Sawbridge, Tim, Spangenberg, German, Webb, Steven R., Gupta, Manju, Ainley, William Michael, Henry, Matthew J., Miller, Jeffrey C., Guschin, Dmitry Y.
• Primary Examiner(s): Keogh, Matthew R
• Application Number:
  US14/019,244
• Publication Number:
  US 20140090112A1
• Time in Patent Office: 1,706 Days
• Field of Search: None
• CPC Class Codes: A01H 5/10 Seeds
• CPC Class Codes: A01H 6/542 Glycine max [soybean]
• CPC Class Codes: C12N 15/102 Mutagenizing nucleic acids
• CPC Class Codes: C12N 15/8216 Methods for controlling, regulating or enhancing expression of transgenes in plant cells
• CPC Class Codes: C12N 15/8247 involving modified lipid metabolism, e.g. seed oil composition
• CPC Class Codes: C12N 15/902 using homologous recombination
• CPC Class Codes: C12N 9/22 Ribonucleases RNAses, DNAses catalytic nucleic acids C12N15/113
• CPC Class Codes: C12Y 301/00 Hydrolases acting on ester bonds (3.1)
• CPC Class Codes: Y02A 40/146 Genetically Modified [GMO] plants, e.g. transgenic plants

Abstract

A method of gene editing or gene stacking within a FAD2 loci by cleaving, in a site directed manner, a location in a FAD2 gene in a cell, to generate a break in the FAD2 gene and then ligating into the break a nucleic acid molecule associated with one or more traits of interest is disclosed.

Claims

1. A transgenic cell comprising a non-naturally occurring exogenous nucleotide sequence integrated within any of SEQ ID Nos:

17-26, 28-33 or 35-38 into an endogenous FAD2A, FAD2A′

, FAD2C and/or FAD2C′

gene, wherein the non-naturally occurring exogenous nucleotide sequence comprises non-coding sequences comprising a plurality of unique nuclease target sites and two or more expression cassettes separated by the non-coding sequences.

2. The transgenic cell of claim 1, further comprising a donor molecule comprising a sequence flanked by regions of homology to the integrated exogenous nucleotide sequence.

3. The transgenic cell of claim 1, wherein the exogenous nucleotide sequence is integrated into some but not all copies of the FAD2A, FAD2A′

, FAD2C and/or FAD2C′

gene.

4. A transgenic plant or seed, comprising the transgenic cell of claim 1.

5. The transgenic cell of claim 1, wherein presence of the exogenous nucleotide sequence in the FAD2A, FAD2A′

, FAD2C and/or FAD2C′

gene does not negatively influence the agronomic or quality properties of the plant.