CRISPR PAM Sequence Table
A PAM sequence is the short DNA motif a CRISPR nuclease must see next to a target before it can cut. This table lists common nucleases, their PAMs, guide spacer lengths, PAM orientation and typical cut-position rules for guide RNA design.
| Nuclease | PAM | Orientation | Spacer | Cut rule | Notes |
|---|---|---|---|---|---|
SpCas9 Streptococcus pyogenes | NGG | 3' of spacer | 20 nt | ~3 bp upstream of PAM | Most common Cas9 system; broad tool and protocol support. |
SpCas9-NG Engineered SpCas9 variant | NG | 3' of spacer | 20 nt | ~3 bp upstream of PAM | Relaxed PAM requirement; useful when NGG sites are scarce. |
SaCas9 Staphylococcus aureus | NNGRRT | 3' of spacer | 21 nt | ~3 bp upstream of PAM | Smaller Cas9 ortholog, often useful for AAV delivery. |
Cas12a / Cpf1 AsCas12a or LbCas12a | TTTV | 5' of spacer | 23-24 nt | Staggered cut distal to PAM | T-rich PAM; creates sticky ends instead of blunt Cas9 cuts. |
Reading PAM ambiguity codes
PAMs use the same IUPAC nucleotide ambiguity codes as sequence motifs. For example, NGG means any base followed by two G bases, and TTTV means TTT followed by A, C or G.
| Code | Meaning |
|---|---|
| N | A, C, G or T (any base) |
| R | A or G (purine) |
| V | A, C or G (not T) |
Frequently asked questions
- What is a PAM sequence?
- A PAM (protospacer-adjacent motif) is a short DNA sequence next to the target site that a CRISPR nuclease must recognise before it can bind and cut. Without the right PAM, the guide RNA will not target that site efficiently.
- What PAM does SpCas9 use?
- The standard Streptococcus pyogenes Cas9 (SpCas9) recognises an NGG PAM on the genomic DNA, where N can be any base. The 20 nt guide spacer sits immediately upstream of the PAM.
- Is the PAM part of the guide RNA?
- No. The guide RNA spacer matches the protospacer sequence next to the PAM, but the PAM itself is present in the target DNA and is not included in the guide RNA.
- How is Cas12a different from Cas9?
- Cas12a recognises a T-rich PAM on the 5' side of the spacer and makes staggered cuts. Cas9 PAMs are usually on the 3' side of the spacer and SpCas9 makes a blunt cut near the PAM.
Related tools and references
Use these related pages when this table raises a practical calculation or workflow question.