摘要
根据大肠杆菌W3110菌株的waaL基因序列设计CRISPR/Cas9的作用靶点,构建sgRNA表达质粒,然后设计同源修复供体DNA序列,通过电转化法导入宿主菌内,从而构建完整的大肠杆菌CRISPR/Cas9基因编辑系统。结果显示,应用该系统成功地构建了大肠杆菌waaL基因缺失株。将该系统继续用于大肠杆菌W3110菌株wecA基因的缺失,结果表明,该系统可快速高效地用于大肠杆菌基因缺失,这为构建大肠杆菌生物工程菌奠定基础。
The binding site of CRISPR/Cas9 was designed based on the waaL gene sequence of the Escherichia coli(E.coli)W3110 strain,and the sgRNA expression plasmid was constructed.The homologous repairing sequence(donor DNA)was designed and introduced into the host bacteria by electrotransformation to construct a complete E.coli CRISPR/Cas9 gene editing system.The results showed that the waaL gene deletion strain of E.coli W3110 was successfully constructed using this system.The system was examined again by the deletion of wecA gene of E.coli W3110 strain.The results showed that this system can be used for E.coli gene deletion quickly and efficiently,which lays the foundation for the construction of engineering E.coli.
作者
卞晓萍
刘青
孔庆科
罗洪艳
李沛
BIAN Xiaoping;LIU Qing;KONG Qingke;LUO Hongyan;LI Pei(College of Animal Science and Technology,South-west University,Chongqing 400700,China)
出处
《中国兽医学报》
CAS
CSCD
北大核心
2021年第1期110-116,共7页
Chinese Journal of Veterinary Science
基金
中央高校基本科研业务费专项资金资助项目(SWU118073,SWU118074)
重庆市自然科学基金资助项目(cstc2019jcyj-msxmX0532)。
