CRISPR enzymes and systems
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. Methods for making and using and uses of such systems, methods, and compositions and products from such methods and uses are also disclosed and claimed.
1. A method for detecting a target DNA in a eukaryotic nucleic acid sample, comprising:
(a) contacting the nucleic acid sample with a CRISPR-Cpf1 complex comprising a Cpf1 effector protein and a guide polynucleotide that comprises a guide sequence linked to a direct repeat sequence, wherein the guide sequence directs sequence-specific binding of the CRISPR-Cpf1 complex to the target DNA, and wherein the complex lacks a tracr sequence; and
(b) detecting presence of the target DNA by detecting occurrence of binding of the CRISPR-Cpf1 complex to the target DNA or cleavage of the target DNA by the CRISPR-Cpf1 complex.
2. The method of claim 1 , wherein the Cpf1 effector protein is of a bacterial species selected from the group consisting of Francisella tularensis 1 , Francisella tularensis subsp. novicida, Prevotella albensis, Lachnospiraceae bacterium MC2017 1 , Butyrivibrio proteoclasticus, Peregrinibacteria bacterium GW2011_GWA2_33_10 , Parcubacteria bacterium GW2011_GWC2_44_17 , Smithella sp. SCADC, Acidaminococcus sp. BV3L6 , Lachnospiraceae bacterium MA2020 , Candidatus Methanoplasma termitum, Eubacterium eligens, Moraxella bovoculi 237 , Leptospira inadai, Lachnospiraceae bacterium ND2006 , Porphyromonas crevioricanis 3 , Prevotella disiens and Porphyromonas macacae.
3. The method of claim 1 , wherein the Cpf1 effector protein is Acidaminococcus sp. BV3L6 Cpf1 (AsCpf1), Lachnospiraceae bacterium ND2006 Cpf1 (LbCpf1), or Franscisella novicida U112 Cpf1 (FnCpf1).
4. The method of claim 3 , wherein the Cpf1 effector protein is Franscisella novicida U112 Cpf1.
5. The method of claim 3 , wherein the Cpf1 effector protein is Acidaminococcus sp. BV3L6 Cpf1.
6. The method of claim 3 , wherein the Cpf1 effector protein is Lachnospiraceae bacterium ND2006 Cpf1.
7. The method of claim 1 , wherein the occurrence of binding of the CRISPR-Cpf1 complex to the target DNA or cleavage of the target DNA by the CRISPR-Cpf1 complex is detected by a fluorescent signal.
8. The method of claim 1 , wherein the target DNA is associated with a disease.
9. The method of claim 1 , wherein the target DNA is associated with cancer.
10. The method of claim 1 , wherein the target DNA is associated with a virus.
11. The method of claim 1 , wherein the target DNA comprises a target sequence hybridizable with the guide sequence, wherein the target sequence is located at 3′ of a Protospacer Adjacent Motif (PAM).
12. The method of claim 11 , wherein the PAM comprises a 5′ T-rich motif.
13. The method of claim 11 , wherein the PAM sequence is TTN, where N is A/C/G or T and the effector protein is FnCpf1, or wherein the PAM sequence is TTTV, where V is A/C or G and the effector protein is PaCpf1, LbCpf1 or AsCpf1.
14. The method of claim 1 , wherein the nucleic acid sample is obtained from a human subject.
15. The method of claim 1 , wherein the nucleic acid sample is contacted with the CRISPR-Cpf1 complex in vitro.