Department of Biochemical Engineering and Biotechnology, DBT-AIST International Laboratory for Advanced Biomedicine (DAILAB), Indian Institute of Technology Delhi, Hauz Khas, New Delhi, India.
Methods Mol Biol. 2022;2349:147-166. doi: 10.1007/978-1-0716-1585-0_8.
The easily programmable CRISPR/Cas9 system has found applications in biomedical research as well as microbial and crop applications, due to its ability to create site-specific edits. This powerful and flexible system has also been modified to enable inducible gene regulation, epigenome modifications and high-throughput screens. Designing efficient and specific guides for the nuclease is a key step and also a major challenge in effective application. This chapter describes rules for sgRNA design and important features to consider while touching upon bioinformatics advances in predicting efficient guides. Computational tools that suggest improved guides, depending on application, or predict off-targets have also been mentioned and compared.
易于编程的 CRISPR/Cas9 系统因其能够进行定点编辑，已在生物医学研究以及微生物和作物应用中得到应用。该强大且灵活的系统还经过了修改，以实现诱导型基因调控、表观基因组修饰和高通量筛选。设计有效的、针对特定靶点的向导 RNA 是关键步骤，也是有效应用的主要挑战。本章描述了 sgRNA 设计规则和在涉及预测有效向导的生物信息学进展时需要考虑的重要特征。还提到并比较了根据应用情况建议改进的向导或预测脱靶的计算工具。
