Composition and methods for regulated expression of a guide rna/cas endonuclease complex
AssigneePioneer Hi-Bred International, Inc.
InventorsANDREW MARK CIGAN, SERGEI SVITASHEV
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Abstract
Compositions and methods are provided for regulated expression of a guide RNA/Cas endonuclease system in a plant cell, plant and seed. Compositions and methods are also provided for genome modification of a target sequence in the genome of a plant or plant cell. The methods and compositions employ a regulated guide RNA/Cas endonuclease system to provide an effective system for modifying or altering target sites within the genome of a plant, plant cell or seed.
Claims20
A method for regulated expression of a guide RNA/Cas endonuclease complex in a plant cell, the method comprising:
- a. providing a guide RNA to a plant cell which has stably incorporated in its genome a recombinant DNA construct comprising an inducible promoter operately linked to a Cas endonuclease, wherein said guide RNA and Cas endonuclease are capable of forming a complex that enables the Cas endonuclease to introduce a double strand break at a target site in the genome of said plant cell;
- and,
- b. induction of the inducible promoter by chemical or stress treatment on the plant cell, wherein said induction results in the expression of the Cas endonuclease;
A method for modifying a target DNA sequence in the genome of a plant cell, the method comprising:
- a) providing a plant cell which has stably incorporated in its genome a recombinant DNA construct comprising an inducible promoter operately linked to a Cas endonuclease;
- b) providing to the plant cell of (a) a guide RNA, wherein said guide RNA and Cas endonuclease are capable of forming a complex that enables the Cas endonuclease to introduce a double strand break at said target DNA sequence;
- and,
- c) induction of the inducible promoter by chemical or stress treatment on the plant cell of (b), wherein said induction results in the expression of the Cas endonuclease of (a).
A method for altering expression of at least one polynucleotide of interest in a plant cell, the method comprising:
- a) providing a plant cell which has stably incorporated in its genome a recombinant DNA construct comprising an inducible promoter operately linked to a Cas endonuclease;
- b) providing to the plant cell of (a) at least one guide RNA, wherein said guide RNA and Cas endonuclease are capable of forming a complex that enables the Cas endonuclease to introduce a double strand break at said target DNA sequence;
- c) induction of the inducible promoter by chemical or stress treatment on the plant cell of (b), wherein said induction results in the expression of the Cas endonuclease of (a);
- and,
- d) selecting a plant cell wherein the expression of the at least one polynucleotide of interest is increased or decreased.
Dependencies are inferred from the claim text.