Biomedical Informatics & Genomics Center, Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, P.R. China.
Research Institute of Xi'an Jiaotong University, Zhejiang, China.
Brief Bioinform. 2022 Sep 20;23(5). doi: 10.1093/bib/bbac183.
Clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 gene editing technology has been widely used to facilitate efficient genome editing. Current popular sgRNA design tools only consider the sgRNA perfectly matched to the target site and provide the results without any on-target mismatch. We suppose taking on-target gRNA-DNA mismatches into consideration might provide better sgRNA with similar binding activity and reduced off-target sites. Here, we trained a seq2seq-attention model with feedback-loop architecture, to automatically generate sgRNAs with on-target mismatches. Dual-luciferase reporter experiment showed that multiple sgRNAs with three mismatches could achieve the 80% of the relative activity of the perfect matched sgRNA. Meanwhile, it could reduce the number of off-target sites using sgRNAs with on-target mismatches. Finally, we provided a freely accessible web server sgRNA design tool named ExsgRNA. Users could submit their target sequence to this server and get optimal sgRNAs with less off-targets and similar on-target activity compared with the perfect-matched sgRNA.
成簇规律间隔短回文重复（CRISPR）/Cas9 基因编辑技术已被广泛用于促进高效的基因组编辑。目前流行的 sgRNA 设计工具仅考虑与靶位点完全匹配的 sgRNA，并提供没有任何靶位错配的结果。我们假设考虑靶位 gRNA-DNA 错配可能会提供具有相似结合活性和减少脱靶位点的更好 sgRNA。在这里，我们训练了一个具有反馈循环结构的 seq2seq-attention 模型，以自动生成具有靶位错配的 sgRNA。双荧光素酶报告基因实验表明，具有三个错配的多个 sgRNA 可以达到完美匹配 sgRNA 相对活性的 80%。同时，使用具有靶位错配的 sgRNA 可以减少脱靶位点的数量。最后，我们提供了一个免费的 sgRNA 设计工具名为 ExsgRNA 的 Web 服务器。用户可以将他们的靶序列提交给这个服务器，并获得与完美匹配 sgRNA 相比，脱靶较少且靶位活性相似的最优 sgRNA。
