Guide RNAs with chemical modification for prime editing
AssigneeAgilent Technologies, Inc.
InventorsDaniel E. Ryan, Robert Kaiser, Douglas J. Dellinger
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Abstract
Provided herein are compositions and methods for inducing CRISPR/Cas-based editing of a target nucleic acid (e.g., target DNA or target RNA) in vitro or in a cell, using modified prime editing guide RNAs (pegRNAs) that incorporate one or more chemically-modified nucleotides. The modified pegRNAs disclosed herein may be used to induce Cas-mediated incorporation of one or more nucleotide changes and/or targeted mutagenesis of a target nucleic acid. The nucleotide change can include, e.g., one or more nucleotide changes, an insertion of one or more nucleotides, or a deletion of one or more nucleotides.
Claims23
A prime-editing guide RNA (pegRNA) comprising:
- a guide sequence that is capable of hybridizing to a target sequence in a target region of a nucleic acid;
- a segment that interacts with a Cas protein;
- a primer-binding site segment that can bind to the target region;
- and a reverse transcriptase template segment including a sequence with desired edit(s) to the target region;
- wherein the pegRNA comprises a 5′ end and a 3′ end, one of which is a prime-editing end and the other is referred to as a distal end, and
- the pegRNA comprises at least two modified nucleotides within 5 nucleotides of the prime-editing end, said at least two modified nucleotides comprising 2′-O-methyl-3′-phosphonoacetate (“MP”).
Dependencies are inferred from the claim text.