CPC C12N 15/82 (2013.01) [C12N 9/22 (2013.01); C12N 15/113 (2013.01); C12N 2310/20 (2017.05)] | 15 Claims |
1. A method of modifying a target nucleic acid, the method comprising:
contacting the target nucleic acid with
(a) a Type V CRISPR-Cas effector protein;
(b) a reverse transcriptase; and
(c) an extended guide nucleic acid, wherein the extended guide nucleic acid comprises:
(i) a Type V CRISPR nucleic acid; and
(ii) an extended portion comprising a primer binding site and a reverse transcriptase template (RT template),
wherein the extended portion of the extended guide nucleic acid comprises an RT template and a primer binding site, optionally wherein the extended portion is fused to either the 5′ end or 3′ end of the Type V CRISPR nucleic acid and/or the RT template of the extended portion is 5′ of the primer binding site, and
wherein the target nucleic acid is double stranded comprising a first strand and a second strand, the Type V CRISPR-Cas effector protein is a double stranded nuclease that cuts the first strand and the second strand of the target nucleic acid resulting in a double stranded break, and the primer binding site binds to the first strand of the target nucleic acid, which is the target strand and same strand to which the Type V CRISPR-Cas effector protein is recruited, thereby modifying the target nucleic acid.
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