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co-crispr: A valuable toolkit for mutation enrichment in the gene editing of Spodoptera frugiperda

Insect Science 2023年第3期30卷 625-636页

作者： Wei-Kang Han Feng-Xian Tang Hao-Li Gao Yan Wang Na Yu Jian-Jun Jiang Ze-Wen Liu Key Laboratory of Integrated Management of Crop Diseases and Pests(Ministry of Education) College of Plant ProtectionNanjing Agricultural UniversityNanjingChina Plant Protection Research Institute Guangxi Academy of Agricultural ScienceGuangxi Key Laboratory of Biology for Crop Diseases and Insect PestsNanningChina

The crispr/Cas9 system has been successfully applied in dozens of diverse species;although the screening of successful crispr/Cas9 editing events remains particularly laborious,especially for those that occur at relatively low ***,a co-crispr strategy was proved to enrich the desired crispr ***,the co-crispr strategy was developed in the Fall armyworm,Spodoptera frugiperda,with kynurenine 3-monooxygenase gene(kmo)as a *** kmo mosaics induced by single-guide RNAs(sgRNAs)/Cas9 displayed the darker green color phenotype in larvae,compared with wild type(brown),and mosaic-eye adults were significantly acquired from the mosaic larvae *** the kmo knockout strain,no significant difference was observed in larval development and adult *** 2(ace2)and Wntl were selected as target genes to construct the co-crispr strategy using kmo *** coinjection of kmo and ace2 sgRNAs,the mutant efficiency of ace2 was significantly increased in the kmo mosaic(larvae or adults)***,more malformed pupae with Wntl mutations were observed in the darker green larvae *** together,these results demonstrated that kmo was a suitable visible marker gene for the application and extension of co-crispr strategy in Fall *** darker green color in larvae or mosaic-eye in adults from kmo knockout as a marker,the mutant efficiency of a target gene could be enriched in a Fall armyworm group consisting of marked *** co-crispr strategy is helpful for gene function studies by the knockout technique with no or lethal phenotypes.

关键词： co-crispr kmo mutation enrichment Spodoptera frugiperda Wnt1

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