Novel cas systems and methods of use

A method for modifying a target site in the genome of a cell, the method comprising introducing into said cell at least one chimeric engineered guide RNA, and a Cas endonuclease comprising a first nuclease domain of SEQ ID NO:

• 88, and a second nuclease domain comprising at least one nuclease sub-domain selected from the group consisting of SEQ ID NO:
• 90, SEQ ID NO:
• 92 and SEQ ID NO:
• 94, wherein said chimeric engineered guide RNA and Cas endonuclease can form a complex that is capable of recognizing, binding to, and optionally nicking, cleaving, or covalently attaching to all or part of said target site;
• further comprising identifying at least one cell that has a modification at said target, wherein the modification at said target site is selected from the group consisting of (i) a replacement of at least one nucleotide, (ii) a deletion of at least one nucleotide, (iii) an insertion of at least one nucleotide, and (iv) any combination of (i)-(iii).

A method for editing a nucleotide sequence in the genome of a cell, the method comprising introducing into said cell at least one polynucleotide modification template, at least one chimeric engineered guide RNA, and a Cas endonuclease comprising a first nuclease domain of SEQ ID NO:

• 88, and a second nuclease domain comprising at least one nuclease sub-domain selected from the group consisting of SEQ ID NO:
• 90, SEQ ID NO:
• 92 and SEQ ID NO:
• 94, wherein said polynucleotide modification template comprises at least one nucleotide modification of said nucleotide sequence, wherein said chimeric engineered guide RNA and Cas endonuclease can form a complex that is capable of recognizing, binding to, and optionally nicking, cleaving, or covalently attaching to all or part of said target site.

A method for modifying a target site in the genome of a cell, the method comprising providing to said cell at least one chimeric engineered guide RNA, at least one donor DNA, and a Cas endonuclease comprising a first nuclease domain of SEQ ID NO:

• 88, and a second nuclease domain comprising at least one nuclease sub-domain selected from the group consisting of SEQ ID NO:
• 90, SEQ ID NO:
• 92 and SEQ ID NO:
• 94, wherein said at least one chimeric engineered guide RNA and Cas endonuclease can form a complex that is capable of recognizing, binding to, and optionally nicking, cleaving, or covalently attaching to all or part of said target site, wherein said donor DNA comprises a polynucleotide of interest;
• further comprising identifying at least one cell that said polynucleotide of interest integrated in or near said target site.

A recombinant DNA polynucleotide comprising a promoter operably linked to a plant-optimized polynucleotide encoding a Cas endonuclease, wherein said Cas endonuclease comprises a first nuclease domain of SEQ ID NO:

• 88, and a second nuclease domain comprising at least one nuclease sub-domain selected from the group consisting of SEQ ID NO:
• 90, SEQ ID NO:
• 92 and SEQ ID NO:
• 94.

A kit for binding, cleaving or nicking a target sequence in a prokaryotic or eukaryotic cell or organism, said kit comprising a guide polynucleotide specific for said target sequence, and a Cas endonuclease or a polynucleotide encoding said Cas endonuclease, wherein said Cas endonuclease comprises a first nuclease domain of SEQ ID NO:

• 88, and a second nuclease domain comprising at least one nuclease sub-domain selected from the group consisting of SEQ ID NO:
• 90, SEQ ID NO:
• 92 and SEQ ID NO:
• 94, wherein said guide polynucleotide is capable of forming a guide polynucleotide/Cas endonuclease complex, wherein said complex can recognize, bind to, and optionally nick or cleave said target sequence.