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 having at least 95% identity to the nucleotide sequence of SEQ ID No: 24 or SEQ ID No: 44, wherein the nucleic acid encodes a nucleic acid-guided nuclease comprising the amino acid sequence of SEQ ID No. 4;
(b) an engineered guide nucleic acid sequence capable of complexing with the nucleic acid-guided nuclease, wherein the engineered guide nucleic acid comprises any one of SEQ ID No. 84, SEQ ID No. 85, SEQ ID No. 87, SEQ ID No. 93, SEQ ID No. 94, SEQ ID No. 95 and SEQ ID No. 96;
(c) an editing sequence having a change in sequence relative to the sequence of a target region in a genome of a cell and a mutation in a protospacer adjacent motif (PAM);
wherein the system results in a genome edit in the target region in the genome of the cell facilitated by the nuclease, the engineered guide nucleic acid, and the editing sequence.
2. The system of claim 1 , wherein the engineered guide nucleic acid and the editing sequence are provided as a single nucleic acid.
3. The system of claim 1 , wherein the nucleic acid encoding the nucleic acid-guided nuclease is codon optimized for Escherichia coli.
4. The system of claim 1 , wherein the nucleic acid encoding the nucleic acid-guided nuclease is codon optimized for Saccharomyces cerevisiae.
5. The system of claim 1 , wherein the nucleic acid encoding the nucleic acid-guided nuclease is codon optimized for mammalian cells.
6. The system of claim 1 , wherein the engineered guide nucleic acid capable of complexing with the nucleic acid-guided nuclease comprises the nucleotide sequence of SEQ ID No. 85.
7. The system of claim 1 , wherein the engineered guide nucleic acid capable of complexing with the nucleic acid-guided nuclease comprises the nucleotide sequence of SEQ ID No: 84.
8. The system of claim 1 , wherein the engineered guide nucleic acid capable of complexing with the nucleic acid-guided nuclease comprises the nucleotide sequence of SEQ ID No. 87.
9. The system of claim 1 , wherein the engineered guide nucleic acid capable of complexing with the nucleic acid-guided nuclease comprises the nucleotide sequence of SEQ ID No. 93.
10. The system of claim 1 , wherein the engineered guide nucleic acid capable of complexing with the nucleic acid-guided nuclease comprises the nucleotide sequence of SEQ ID No. 94.
11. The system of claim 1 , wherein the engineered guide nucleic acid capable of complexing with the nucleic acid-guided nuclease comprises the nucleotide sequence of SEQ ID No. 95.
12. The system of claim 1 , wherein the engineered guide nucleic acid capable of complexing with the nucleic acid-guided nuclease comprises the nucleotide sequence of SEQ ID No. 96.
13. The system of claim 1 , wherein the target region is within a eukaryotic cell.
14. The system of claim 1 , wherein the target region is within a plant cell.
15. The system of claim 1 , wherein the target region is within a bacterial cell.
16. The system of claim 1 , wherein the target region is within a mammalian cell.
17. The system of claim 1 , wherein the target region is within a coding region of a protein.
18. The system of claim 1 , wherein the target region is within a non-coding region of a protein.
19. The system of claim 18 , wherein the target region is within a regulatory region of a protein.
20. A nucleic acid nuclease having at least 95% identity to the nucleotide sequence of SEQ ID No: 44.