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US 9,885,033 B2Granted
Increasing specificity for RNA-guided genome editing
AssigneeThe General Hospital Corporation
InventorsJ. Keith Joung, James Angstman, Jeffry D. Sander, Morgan Maeder, Shengdar Tsai
Life of this patent
Estimating the term…
Abstract
Methods for increasing specificity of RNA-guided genome editing, e.g., editing using CRISPR/Cas9 systems.
Claims3
A method of inducing a single or double-stranded break in a target region of a double-stranded DNA molecule in a cell, the method comprising expressing in or introducing into the cell:
- a Streptococcus pyogenes Cas9 nuclease or nickase;
- and
- a guide RNA that consists of a sequence selected from the group consisting of:
- (SEQ ID NO:
- 4)
- (X17-20)GUUUUAGAGCUAUGCUGUUUUG(XN);
- (SEQ ID NO:
- 5)
- (X17-20)GUUUUAGAGCUA;
- (SEQ ID NO:
- 6)
- (X17-20)GUUUUAGAGCUAUGCUGUUUUG;
- (SEQ ID NO:
- 7)
- (X17-20)GUUUUAGAGCUAUGCU;
- (SEQ ID NO:
- 8)
- (X17-20)GUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCG
- (XN);
- (SEQ ID NO:
- 9)
- (X17-20)GUUUUAGAGCUAUGCUGAAAAGCAUAGCAAGUUAAAAUAAGG
- CUAGUCCGUUAUC(XN);
- (SEQ ID NO:
- 10)
- (X17-20)GUUUUAGAGCUAUGCUGUUUUGGAAACAAAACAGCAUAGCAA
- GUUAAAAUAAGGCUAGUCCGUUAUC(XN);
- (SEQ ID NO:
- 11)
- (X17-20)GUUUUAGAGCUAGAAAUAGCAAGUUAAAAUAAGGCUAGUCCG
- UUAUCAACUUGAAAAAGUGGCACCGAGUCGGUGC(XN),
- (SEQ ID NO:
- 12)
- (X17-20)GUUUAAGAGCUAGAAAUAGCAAGUUUAAAUAAGGCUAGUCCG
- UUAUCAACUUGAAAAAGUGGCACCGAGUCGGUGC;
- (SEQ ID NO:
- 13)
- (X17-20)GUUUUAGAGCUAUGCUGGAAACAGCAUAGCAAGUUUAAAUAA
- GGCUAGUCCGUUAUCAACUUGAAAAAGUGGCACCGAGUCGGUGC;
- or
- (SEQ ID NO:
- 14)
- (X17-20)GUUUAAGAGCUAUGCUGGAAACAGCAUAGCAAGUUUAAAUAA
- GGCUAGUCCGUUAUCAACUUGAAAAAGUGGCACCGAGUCGGUGC;
- wherein X17-20 is a sequence complementary to 17-20 consecutive nucleotides of the complementary strand of a target sequence, preferably a target sequence immediately 5′ of a protospacer adjacent motif (PAM);
- N is 0;
- and:
- the guide RNA includes one or more deoxyribonucleotides.
45 more elements
Dependencies are inferred from the claim text.