The CRISPR/Cas9 system is an RNA-guided endonuclease technology for gene editing. This system requires guide RNA (gRNA) comprising a 20 bp target sequence adjacent to a PAM (Protospacer Adjacent Motif) site to direct the Cas9 endonuclease to the cleavage site. The Find CRISPR sites tool in Geneious will search for gRNA ("CRISPR") sites in the selected gene, and search for off-target binding sites in your genome of interest. Each CRISPR site in the selected sequence is scored according to how many off-target sites it potentially will bind to, and how similar the off-target sites are to the original sequence.
In this tutorial you will be working with the LYP1 CDS from Saccharomyces cerevisiae S288c and using the S288c genome as the off-target database. LYP1, or Lysine permease, is found on chromosome 14 in the yeast genome. It is one of three amino acid permeases (Alp1p, Can1p, Lyp1p) responsible for uptake of cationic amino acids. For more information about this gene, click here.