CRISPR-SE: a brute force search engine for CRISPR design.

Abstract

CRISPR is a revolutionary genome-editing tool that has been broadly used and integrated within novel biotechnologies. A major component of existing CRISPR design tools is the search engines that find the off-targets up to a predefined number of mismatches. Many CRISPR design tools adapted sequence alignment tools as the search engines to speed up the process. These commonly used alignment tools include BLAST, BLAT, Bowtie, Bowtie2 and BWA. Alignment tools use heuristic algorithm to align large amount of sequences with high performance. However, due to the seed-and-extend algorithms implemented in the sequence alignment tools, these methods are likely to provide incomplete off-targets information for ultra-short sequences, such as 20-bp guide RNAs (gRNA). An incomplete list of off-targets sites may lead to erroneous CRISPR design. To address this problem, we derived four sets of gRNAs to evaluate the accuracy of existing search engines; further, we introduce a search engine, namely CRISPR-SE. CRISPR-SE is an accurate and fast search engine using a brute force approach. In CRISPR-SE, all gRNAs are virtually compared with query gRNA, therefore, the accuracies are guaranteed. We performed the accuracy benchmark with multiple search engines. The results show that as expected, alignment tools reported an incomplete and varied list of off-target sites. CRISPR-SE performs well in both accuracy and speed. CRISPR-SE will improve the quality of CRISPR design as an accurate high-performance search engine.