CRISPR guide off-target check
crispr_offtarget_checkScreen a guide's protospacer for off-target sites (protospacer match + valid PAM, both strands) against a small curated set of common lab reference genomes (see genomesChecked) — NOT a whole human/mouse genome search. For SpCas9 with a 20 nt spacer each site also gets a Doench 2016 CFD score, so sites are ranked by predicted cut likelihood rather than by mismatch count alone, and the guide gets an aggregate specificity. Use this the same way primer_specificity is used: a useful sanity check within the covered organisms, not a clearance guarantee for a mammalian expression host. PREDICTED, NOT MEASURED. Best of the common off-target scores on the authors' GUIDE-Seq comparison, at Pearson r = 0.40 over 9 guides and 402 sites (vs CCTop 0.31, Hsu-Zhang 0.26) — a useful ranking, not a reliable magnitude. Weights were measured for SINGLE mismatches; multiple mismatches are multiplied, and Listgarten et al. 2018 note the training data never contained a mismatch and an alternative PAM together, so that combination is extrapolation. Valid for: SpCas9 with a 20 nt spacer, which is the only case scored — every other nuclease returns null rather than a number from the wrong enzyme's table. One locus, one cell line. Substitutions only: DNA/RNA bulges are neither searched nor scorable, and a low score is not a claim that a site is safe.
Input Schema
| Name | Required | Description | Default |
|---|---|---|---|
| nuclease | No | Nuclease id — determines the PAM pattern/side required at each candidate site. | spcas9 |
| protospacer | Yes | The guide's protospacer sequence, 5'→3' (no PAM). Max 32 nt — every supported nuclease uses a 20–23 nt guide. | |
| maxMismatches | No | Mismatches tolerated between the protospacer and a candidate genomic site. Max 4 — a complete search seeds on maxMismatches+1 non-overlapping blocks, and past that the blocks are too short to be selective against a multi-Mb genome (a site that mismatched more would not be cut anyway). |