厦门大学学报（自然科学版）2016，Vol.55Issue(6)：922-926,5.DOI:10.6043/j.issn.0438-0479.201604028
利用CRISPR/Cas9技术制备Mindin基因敲除小鼠
Application of CRISPR/Cas9 Technology for Establishing Mindin Gene Knockout Mice
摘要
Abstract
To establish Mindin knockout mice using CRISPR/Cas9 gene targeting technology,three sgRNA targets of Mindin were designed according to Mindin genome sequence.Vectors expressing sgRNA and Cas9 were constructed separately,and co-transfected into L929 cells by with electroporation method.Transfected cells were selected with blasticitin and the mutation rate was determined using DNA sequencing with proper primers.The sgRNA which had resulted in the highest mutation rate in L929 cells was trans-cribed in vitro,along with Cas9.Then sgRNA and Cas9 mRNAs were microinjected into mice zygotes and 40 mice were born after the microinj ection.Genotypes of the mice were determined by PCR with genomic DNA from the new born mice tail and 1 7 neonatal mice were found carrying mutations in Mindin gene by subsequent sequencing.Gene sequencing results showed that all mutant mice had different levels of nucleotide insertion or deletion,with the maximum number of 10 base deletion found in mouse No.23,leading to the reading frame shift and Mindin gene silence.Taking together,we have successfully established a mouse model with Mindin gene knockout,which lays a solid foundation for related future research.关键词
CRISPR/Cas9/基因敲除/MindinKey words
CRISPR/Cas9/gene knockout/Mindin分类
生物科学引用本文复制引用
纪慈数,赵志刚,贺颖..利用CRISPR/Cas9技术制备Mindin基因敲除小鼠[J].厦门大学学报（自然科学版）,2016,55(6):922-926,5.基金项目
福建省自然科学基金重点项目(2013Y01010490) （2013Y01010490）
