SgRNA engineering for improved genome editing and expanded functional assays

[Display omitted] •SgRNA engineering enables more powerful and versatile CRISPR/Cas9 systems.•The sgRNA engineering strategies are summarized in a structural manner.•Optimization of sgRNA sequences leads to better genome editing performance.•Engineering sgRNA with extra modules expands CRISPR-based functional assays. The CRISPR/Cas system has been established as the most powerful and practical genome engineering tool for both fundamental researches and biotechnological applications. Great efforts have been devoted to engineering the CRISPR system with better performance and novel functions. As an essential component, single guide RNAs (sgRNAs) have been extensively designed and engineered with desirable functions. This review highlights representative studies that optimize the sgRNA nucleotide sequences for improved genome editing performance (e.g. activity and specificity) as well as add extra aptamers and end extensions for expanded CRISPR-based functional assays (e.g. transcriptional regulation, genome imaging, and prime editor). The perspectives for further sgRNA engineering to establish more powerful and versatile CRISPR/Cas systems are also discussed.