A CRISPR-Cas9 system is loaded with a guide RNA (gRNA) and incubated with double-stranded DNA. Cas9 introduces a double-strand break adjacent to a 5'-NGG-3' PAM motif.
Which feature determines where Cas9 introduces a double-strand break in target DNA?
- A
The presence of histones at the locus.
- Bcheck_circle
The sequence of the guide RNA, which base-pairs with the target DNA adjacent to a PAM.
- C
DNA methylation status only.
- D
Random scanning of the genome by Cas9 alone.
Explanation
Cas9 is targeted by its guide RNA (sgRNA), which base-pairs with a complementary 20-nt protospacer adjacent to a PAM motif (e.g., NGG). The complementarity dictates editing specificity.