RNA-guided nucleic acid modifying enzymes and methods of use thereof
First Claim
1. A method of modifying a target DNA, the method comprising contacting the target DNA with:
- A) a CasX polypeptide; and
B) a CasX guide RNA comprising a guide sequence that hybridizes to a target sequence of the target DNA,wherein said contacting results in modification of the target DNA by the CasX polypeptide, andwherein said contacting;
a) comprises;
introducing into a cell;
(i) the CasX polypeptide, or a nucleic acid comprising a nucleotide sequence encoding the CasX polypeptide, and (ii) the CasX guide RNA, or a nucleic acid comprising a nucleotide sequence encoding the CasX guide RNA;
orb) takes place in vitro outside of a cell.
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Abstract
The present disclosure provides CasX proteins, nucleic acids encoding the CasX proteins, and modified host cells comprising the CasX proteins and/or nucleic acids encoding same. CasX proteins are useful in a variety of applications, which are provided. The present disclosure provides CasX guide RNAs that bind to and provide sequence specificity to the CasX proteins, nucleic acids encoding the CasX guide RNAs, and modified host cells comprising the CasX guide RNAs and/or nucleic acids encoding same. CasX guide RNAs are useful in a variety of applications, which are provided. The present disclosure provides archaeal Cas9 polypeptides and nucleic acids encoding same, as well as their associated archaeal Cas9 guide RNAs and nucleic acids encoding same.
24 Citations
30 Claims
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1. A method of modifying a target DNA, the method comprising contacting the target DNA with:
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A) a CasX polypeptide; and B) a CasX guide RNA comprising a guide sequence that hybridizes to a target sequence of the target DNA, wherein said contacting results in modification of the target DNA by the CasX polypeptide, and wherein said contacting; a) comprises;
introducing into a cell;
(i) the CasX polypeptide, or a nucleic acid comprising a nucleotide sequence encoding the CasX polypeptide, and (ii) the CasX guide RNA, or a nucleic acid comprising a nucleotide sequence encoding the CasX guide RNA;
orb) takes place in vitro outside of a cell. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29)
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30. A method of modifying a target DNA, the method comprising contacting the target DNA in a eukaryotic cell with:
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A) a CasX polypeptide; and B) a CasX guide RNA comprising a guide sequence that hybridizes to a target sequence of the target DNA, wherein said contacting results in modification of the target DNA by the CasX polypeptide, and wherein said contacting comprises introducing into the eukaryotic cell;
(i) the CasX polypeptide, or a nucleic acid comprising a nucleotide sequence encoding the CasX polypeptide, and (ii) the CasX guide RNA, or a nucleic acid comprising a nucleotide sequence encoding the CasX guide RNA.
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Specification