Nucleic acid-guided nucleases
Disclosed herein are nucleic acid-guided nucleases, guide nucleic acids, and targetable nuclease systems, and methods of use. Disclosed herein are engineered non-naturally occurring nucleic acid-guided nucleases, guide nucleic acids, and targetable nuclease systems, and methods of use. Targetable nuclease systems can be used to edit genetic targets, including recursive genetic engineering and trackable genetic engineering methods.
1. A nucleic acid-guided nuclease system comprising:
(a) a nucleic acid-guided nuclease comprising the amino acid sequence of SEQ ID NO: 7;
(b) a heterologous engineered guide nucleic acid for complexing with the nucleic acid-guided nuclease, wherein said heterologous engineered guide comprises a pseudoknot region comprising nucleotide residues 17-26 of SEQ ID NO: 200 or nucleotide residues 17-26 of SEQ ID NO: 201, and
(c) an editing sequence having a change in sequence relative to the sequence of a target region; wherein the system results in an edit in the target region facilitated by the nuclease, the heterologous guide nucleic acid, and the editing sequence.
2. The system of claim 1 , wherein the target region is within a eukaryotic cell.
3. The system of claim 1 , wherein the target region is within a bacterial cell.
4. The system of claim 1 , wherein the target region is within a plant cell.
5. The system of claim 1 , wherein the target region is within a mammalian cell.
6. The system of claim 1 , wherein the heterologous engineered guide nucleic acid comprises any one of SEQ ID NO: 88, SEQ ID NO: 93, SEQ ID NO: 94, and SEQ ID NO: 95.
7. The system of claim 1 , wherein the heterologous engineered guide nucleic acid and the editing sequence are provided as a single nucleic acid.
8. A method of modifying a target region, the method comprising:
(a) contacting a sample comprising the target region with:
(i) a nucleic acid-guided nuclease comprising the amino acid sequence of SEQ ID NO: 7; and
(ii) a heterologous engineered guide nucleic acid for complexing with the nucleic acid-guided nuclease, wherein said heterologous engineered guide comprises a pseudoknot region having a sequence comprising nucleotide residues 17-26 of SEQ ID NO: 200 or nucleotide residues 17-26 of SEQ ID NO: 201; and
(b) allowing the nuclease and heterologous guide nucleic acid to modify the target region.
9. The method of claim 8 , wherein the sample is further contacted with (iii) an editing sequence having a change in sequence relative to the sequence of the target region.
10. The method of claim 9 , wherein modifying the target results in an edit in the target region facilitated by the nuclease, the heterologous engineered guide nucleic acid, and the editing sequence.
11. The method of claim 9 , wherein the heterologous engineered guide nucleic acid and the editing sequence are provided as a single nucleic acid.
12. The method of claim 9 , wherein the editing sequence further comprises a mutation in a protospacer adjacent motif (PAM) site.
13. The method of claim 8 , wherein the target region is within a eukaryotic cell.
14. The method of claim 8 , wherein the target region is within a bacterial cell.
15. The method of claim 8 , wherein the target region is within a plant cell.
16. The method of claim 8 , wherein the target region is within a mammalian cell.
17. The method of claim 8 , wherein the heterologous engineered guide nucleic acid comprises any one of SEQ ID NO: 88, SEQ ID NO: 93, SEQ ID NO: 94, and SEQ ID NO: 95.