Compositions and methods for generating diversity at targeted nucleic acid sequences
AssigneeMonsanto Technology LLC
InventorsGraeme S. GARVEY, Farhad MOSHIRI
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Abstract
The present disclosure provides methods and kits useful for generating targeted modifications in target nucleic acids using catalytically inactive guided-nucleases in combination with mutagens.
Claims30
A method of inducing a targeted modification in a target nucleic acid molecule, comprising contacting the target nucleic acid molecule with:
- (a) a catalytically inactive guided-nuclease;
- and
- (b) at least one mutagen,
- wherein at least one modification is induced in the target nucleic acid molecule.
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A method of increasing the mutation rate in a targeted region of a nucleic acid molecule, comprising contacting the nucleic acid molecule with:
- (a) a catalytically inactive guided-nuclease;
- (b) at least one guide nucleic acid, wherein the at least one guide nucleic acid forms a complex with the catalytically inactive guided-nuclease, and wherein the at least one guide nucleic acid hybridizes with the target nucleic acid molecule;
- and
- (c) at least one mutagen,
- wherein the target nucleic acid molecule comprises a protospacer adjacent motif (PAM) site, and wherein the mutation rate in the targeted region of the nucleic acid molecule is increased compared to an untargeted nucleic acid molecule.
A method of increasing allelic diversity in a target region of a nucleic acid molecule within a genome of a plant, comprising providing to the plant:
- (a) a catalytically inactive guided-nuclease or a nucleic acid encoding the catalytically inactive guided-nuclease;
- (b) at least one guide nucleic acid or a nucleic acid encoding the at least one guide nucleic acid, wherein the at least one guide nucleic acid forms a complex with the catalytically inactive guided-nuclease, and wherein the at least one guide nucleic acid hybridizes with the nucleic acid molecule;
- and
- (c) at least one mutagen,
- wherein the nucleic acid comprises a protospacer adjacent motif (PAM) site adjacent to the targeted region, and wherein allelic diversity of the target region of the nucleic acid molecule is increased.
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Dependencies are inferred from the claim text.