CrisPam – a tool for designing gRNA sequences to specifically target a variant allele using CRISPR

Abstract

Background & Aim CRISPR is a promising novel technology for treating genetic conditions. Therefore, it is essential to further develop and promote treatment's safety and specificity. While the guide RNA offers position-specific DNA targeting, it may tolerate small changes such as single nucleotide polymorphisms (SNPs). To that end, an allele-specific targeting approach is in need for future treatments of heterozygous patients, suffering from genetic conditions caused by a SNP. The SNP-derived PAM approach allows highly allele-specific DNA cleavage by having a protospacer adjacent motif (PAM) sequence only at the target allele. Here we present CrisPam, a tool that detects SNP-derived PAMs for allele-specific targeting by the CRISPR/Cas system. The CrisPam tool allows researchers to use their SNP data as input as receive an analysis of the DNA sequences revealing the suitable Cas variants, out of 14, for achieving allele-specific targeting. Methods, Results & Conclusion CrisPam is an algorithm that scans DNA sequences of SNPs and detects PAMs generated by pathogenic SNPs. A SNP is accepted as a match if a PAM is found at the pathogenic allele and not at the wildtype allele. Results Using CrisPam we found that 83% of the SNPs generate at least one PAM. 49,826 pathogenic SNPs and 14,755 likely pathogenic SNPs (64,581 in total) were scanned and analyzed. The obtained database contains the SNP information and the Cas proteins for whom a PAM was generated by a pathogenic SNP. Conclusions We have developed CrisPam, a web tool available to researchers for designing their own CRISPR-based experiments for allele-specific targeting. CrisPam offers a SNP-derived PAM solution for targeting pathogenic alleles using the CIRPSR/Cas system.