Skip to main content
. Author manuscript; available in PMC: 2017 Dec 20.
Published in final edited form as: Cold Spring Harb Protoc. 2016 Dec 1;2016(12):pdb.top088849. doi: 10.1101/pdb.top088849

Figure 1.

Figure 1

The three stages of adaptive immunity by Type II-C CRISPR–Cas systems. (A–D) Spacer acquisition: (A) foreign DNA (dark purple) enters the cell, and (B) Cas1, Cas2, and Cas9 in complex with tracrRNA (blue) select a spacer sequence on the target through Cas9-mediated identification of a protospacer adjacent motif (PAM; dark purple rectangle on the foreign DNA). The PAM adjacent sequence is processed into a spacer-sized fragment. (C) The Cas protein complex attached to the spacer identifies the CRISPR array and creates staggered single-stranded breaks on each side on a repeat. (D) The new spacer sequence is inserted into the array and the single-stranded repeats on either side of the new spacer are repaired by DNA polymerase I. (E,F) crRNA transcription and maturation: (E) the CRISPR array and tracrRNA are transcribed. (F) Cas9 binds tracrRNA and the CRISPR transcript, which is then cleaved into mature, spacer-specific crRNAs by RNase III. The mature dual crRNA:tracrRNA remains bound to Cas9 as a hetero-duplex. (G,H) Target identification and cleavage: (G) Upon re-infection with foreign DNA, the spacer on the crRNA of the Cas9:RNA heteroduplex binds to its complementary sequence on the foreign nucleic acid. (H) Cas9 adopts a conformationally active state and cleaves both DNA strands in the target, protecting the cell.