title: CRISPR/Cas9 mediated multiplex genome editing and heritable mutagenesis of BmKu70 in Bombyx mori.
abstract: CRISPR/Cas9, a bacterial adaptive immune system derived genome-editing technique, has become to be one of the most compelling topics in biotechnology. Bombyx mori is an economically important insect and a model organism for studying lepidopteran and arthropod biology. Here we reported highly efficient and multiplex genome editing in B. mori cell line and heritable site-directed mutagenesis of Bmku70, which is required for NHEJ pathway and also related to antigen diversity, telomere length maintenance and subtelomeric gene silencing, using CRISPR/Cas9 system. We established a simple and practicable method and obtained several Bmku70 knockout B. mori lines, and showed that the frequency of HR was increased in embryos of the Bmku70 knockout B. mori. The mutant lines obtained in this study could be a candidate genetic resource for efficient knock-in and fundamental research of DNA repair in B. mori. We also provided a strategy and procedure to perform heritable genome editing of target genes with no significant phenotype effect.
authors.affiliation: 1] State Key Laboratory of Silkworm Genome Biology, Southwest University, Chongqing, 400716 P. R. China [2].
authors.forename: Sanyuan
authors.initials: S
authors.lastname: Ma
authors.affiliation: 1] State Key Laboratory of Silkworm Genome Biology, Southwest University, Chongqing, 400716 P. R. China [2].
authors.forename: Jiasong
authors.initials: J
authors.lastname: Chang
authors.affiliation: State Key Laboratory of Silkworm Genome Biology, Southwest University, Chongqing, 400716 P. R. China.
authors.forename: Xiaogang
authors.initials: X
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Silkworm Genome Biology, Southwest University, Chongqing, 400716 P. R. China.
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Silkworm Genome Biology, Southwest University, Chongqing, 400716 P. R. China.
authors.forename: Jianduo
authors.initials: J
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Silkworm Genome Biology, Southwest University, Chongqing, 400716 P. R. China.
authors.forename: Wei
authors.initials: W
authors.lastname: Lu
authors.affiliation: State Key Laboratory of Silkworm Genome Biology, Southwest University, Chongqing, 400716 P. R. China.
authors.forename: Jie
authors.initials: J
authors.lastname: Gao
authors.affiliation: State Key Laboratory of Silkworm Genome Biology, Southwest University, Chongqing, 400716 P. R. China.
authors.forename: Run
authors.initials: R
authors.lastname: Shi
authors.affiliation: State Key Laboratory of Silkworm Genome Biology, Southwest University, Chongqing, 400716 P. R. China.
authors.forename: Ping
authors.initials: P
authors.lastname: Zhao
authors.affiliation: State Key Laboratory of Silkworm Genome Biology, Southwest University, Chongqing, 400716 P. R. China.
authors.forename: Qingyou
authors.initials: Q
authors.lastname: Xia
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D012831:Bombyx / Q000235:genetics*; D064113:CRISPR-Cas Systems; D002460:Cell Line; D018390:Gene Targeting; D017344:Genes, Insect; D049750:Genome, Insect; D005838:Genotype; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009154:Mutation; D016415:Sequence Alignment