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US 2023/0407341 A1Published application
Using Truncated Guide RNAs (tru-gRNAs) to Increase Specificity for RNA-Guided Genome Editing
AssigneeThe General Hospital Corporation
InventorsJ. Keith Joung, Jeffry D. Sander, Morgan Maeder, Yanfang Fu
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Abstract
Methods for increasing specificity of RNA-guided genome editing, e.g., editing using CRISPR/Cas9 systems, using truncated guide RNAs (tru-gRNAs).
Claims21
-28. (canceled)
A complex comprising:
- a Streptococcus pyogenes Cas9 protein;
- and
- a Streptococcus pyogenes gRNA molecule that includes a complementarity region at the 5′ end of the gRNA consisting of 17-18 nucleotides that are complementary to 17-18 consecutive nucleotides of the complementary strand of a target sequence, wherein the target sequence is immediately 5′ of a protospacer adjacent motif,
- wherein the gRNA is a single gRNA, and
- wherein in the presence of an S. pyogenes Cas9 protein, the gRNA complementary region binds and directs the Cas9 protein to the target sequence.
2 more elements
A complex comprising:
- a Streptococcus pyogenes Cas9 protein;
- and
- a Streptococcus pyogenes gRNA molecule that includes a complementarity region at the end of the gRNA consisting of 17-18 nucleotides that are complementary to 17-18 consecutive nucleotides of the complementary strand of a target sequence, wherein the target sequence is immediately 5′ of a protospacer adjacent motif,
- wherein the gRNA is a CRISPR RNA (crRNA), and
- wherein in the presence of an S. pyogenes Cas9 protein, the gRNA complementary region binds and directs the Cas9 protein to the target sequence.
2 more elements
A vector comprising a DNA molecule encoding:
- a Streptococcus pyogenes gRNA molecule that includes a complementarity region at the end of the gRNA consisting of 17-18 nucleotides that are complementary to 17-18 consecutive nucleotides of the complementary strand of a target sequence, wherein the target sequence is immediately 5′ of a protospacer adjacent motif,
- wherein the gRNA is a single gRNA or a CRISPR RNA (crRNA), and
- wherein in the presence of an S. pyogenes Cas9 nuclease, the gRNA complementary region binds and directs the Cas9 nuclease to the target sequence;
- and
- a promoter.
2 more elements
Dependencies are inferred from the claim text.