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    Circular RNAs (circRNAs) produced from back-spliced exons are widely expressed, but individual circRNA functions remain poorly understood owing to the lack of adequate methods for distinguishing circRNAs from cognate messenger RNAs with overlapping exons. Here, we report that CRISPR-RfxCas13d can effectively discriminate circRNAs from mRNAs by using guide RNAs targeting sequences spanning back-splicing junction (BSJ) sites featured in RNA circles. Using a lentiviral library that targets sequences across BSJ sites of highly expressed human circRNAs, we show that a group of circRNAs are important for cell growth mostly in a cell-type-specific manner and that a common oncogenic circRNA, circFAM120A, promotes cell proliferation by preventing the mRNA for family with sequence similarity 120A (FAM120A) from binding the translation inhibitor IGF2BP2. Further application of RfxCas13d-BSJ-gRNA screening has uncovered circMan1a2, which has regulatory potential in mouse embryo preimplantation development. Together, these results establish CRISPR-RfxCas13d as a useful tool for the discovery and functional study of circRNAs at both individual and large-scale levels.


    Siqi Li, Xiang Li, Wei Xue, Lin Zhang, Liang-Zhong Yang, Shi-Meng Cao, Yun-Ni Lei, Chu-Xiao Liu, Si-Kun Guo, Lin Shan, Man Wu, Xiao Tao, Jia-Lin Zhang, Xiang Gao, Jun Zhang, Jia Wei, Jinsong Li, Li Yang, Ling-Ling Chen. Screening for functional circular RNAs using the CRISPR-Cas13 system. Nature methods. 2021 Jan;18(1):51-59

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    PMID: 33288960

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