Cpf1 complexes with reduced indel activity
AssigneesThe Broad Institute, Inc., Massachusetts Institute of Technology
InventorsFeng ZHANG, Bernd ZETSCHE, Winston YAN, Neville Espi SANJANA, Sara JONES
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Abstract
The invention provides for systems, methods, and compositions for targeting nucleic acids. In particular, the invention provides non-naturally occurring or engineered DNA-targeting systems comprising a novel DNA-targeting CRISPR effector protein and at least one targeting nucleic acid component like a guide RNA.
Claims62
An engineered, non-naturally occurring composition comprising a CRISPR-Cas system, said system comprising a functional CRISPR Cpf1 effector protein and guide RNA (gRNA);
- wherein the gRNA comprises a dead guide sequence;
- whereby the gRNA is capable of hybridizing to a target sequence;
- whereby the CRISPR-Cas system is directed to the target sequence with reduced indel activity resultant from nuclease activity of a non-mutant Cpf1 enzyme of the system.
A cell comprising the non-naturally occurring or engineered composition of any one of the preceding claims.
A method for selecting a guide RNA for directing a CRISPR-Cpf1 system to a target locus in an organism, which comprises:
- a) locating a CRISPR-Cpf1 PAM sequence in the target locus;
- b) selecting a guide sequence that matches a sequence from of 15-23 nucleotides 3′ from the PAM sequence,
- c) determining whether there are off-target genomic sequences that differ from the selected guide sequence by three nucleotides or fewer;
- and
- d) selecting the sequence for use in a guide RNA of a CRISPR-Cpf1 system if the closest off-target sequence differs by one nucleotide or more.
Dependencies are inferred from the claim text.