Figure 1.

Overview of guide RNA biogenesis and genomic targeting by CRISPR-Cas9. Key components of Streptococcus pyogenes CRISPR-Cas locus are shown. Tracr, Cas operon and CRISPR locus (containing clustered repeats and spacers) are transcribed to tracr RNA with stem loop structure, Cas9 endonuclease and pre-crRNA, respectively. RNase III then generate mature crRNA which is eventually incorporated into Cas9 along with tracr RNA. Guide RNA (gRNA) along with RNA-protein complex binds to the specific genomic locus with complementary sequence. Binding of guide RNA and target sequence is facilitated by protospacer adjacent motif (PAM). DNA is cleaved by Cas9 and other accompanying endonucleases thus generating double strand DNA breaks which are eventually repaired by non-homologous end joining (NHEJ) or by homology directed repair (HDR).