CRISPR-Cas9 gene editing uses guide RNA to direct Cas9 nuclease. The PAM (protospacer adjacent motif) sequence required for Cas9 cleavage from Streptococcus pyogenes is:
- A 5'-NGG-3' immediately 3' of the target (non-template strand) ✓
- B 5'-TTTTV-3' on the non-template strand
- C 5'-TGGN-3' immediately 5' of the target
- D 5'-NNNNGATT-3' recognised by Cas12a
Correct answer: A. 5'-NGG-3' immediately 3' of the target (non-template strand)
Explanation
SpCas9 (from S. pyogenes) requires the sequence 5'-NGG-3' (PAM) on the non-template strand immediately 3' of the 20 nt target sequence. The guide RNA (gRNA/sgRNA) base-pairs with the complementary strand; Cas9 causes a blunt-ended double-strand break 3 bp upstream of the PAM. In the absence of PAM, Cas9 does not cleave (PAM sensing prevents self-cleavage of the CRISPR array). PAM requirements vary by Cas orthologue and are being engineered to expand targetable sequences.
Reference: Harper's Illustrated Biochemistry, 32nd ed.
High-yield for: NEET PGINI-CETNExTFMGEUSMLEPLABMRCP
Written and medically reviewed by the StethoPrep medical team.