摘要
以构建pUC18-EGFP载体为例,通过对DNA连接产物进行沉淀处理,对普通的DNA沉淀法进行了改良.实验证明,采用改良的DNA沉淀法处理后的连接产物,转化大肠杆菌的效率约是对照组的10倍,并且大肠杆菌感受态细胞用量减少到10μL.这一改良的DNA沉淀法用CaCl_2替代TE作为溶解沉淀DNA的溶剂,省略了用70%乙醇洗涤DNA沉淀的步骤,较普通的DNA沉淀法操作起来更为简便.
The CaCl2 method is commonly used to transform DNA into Escherichia coli.During construction of recombinant DNA,transformation efficiency of DNA ligation product is rather low.It makes sense to develop new methods to increase transformation efficiency.In this study,for pUC18-EGFP recombinant construction,the DNA precipitation method was improved.Specifically,DNA ligation product was treated with improved DNA precipitation method before transformation,transformation efficiency was ten times of control.It also reduced the use of competent cells down to 10 microliter for recommended quantity.In addition,this improved DNA precipitation replaced TE buffer with calcium chloride to dissolve precipitated DNA and omitted the step of 70%ethanol washing of DNA precipitates.The new method makes manipulation of DNA precipitation method simpler and more convenient.
作者
谭锬
伍金月
唐艺菲
李娜
虞佳
梁前进
TAN Tan;WU Jinyue;TANG Yifei;LI Na;YU Jia;LIANG Qianjin(Hunan Province Cooperative Innovation Center for Molecular Target New Drug Study/Hengyang medical college,University of South China,421001,Hengyang,Hunan,China;Beijing Key Laboratory of Gene Resource and Molecular Development,College of Life Sciences,Beijing Normal University,100875,Beijing,China;Laboratory of Protein Structure and Function,University of South China,421001,Hengyang,Hunan,China)
出处
《北京师范大学学报（自然科学版）》
CAS
CSCD
北大核心
2019年第2期225-232,共8页
Journal of Beijing Normal University(Natural Science)
基金
国家自然科学基金资助项目(31571394)
湖南省分子靶标新药研究协同创新中心开放基金资助项目(0223-0002-0002000-57)
南华大学大学生创新性实验计划资助项目(2017XJYZ041)
关键词
重组DNA
DNA沉淀法
转化
大肠杆菌
recombination DNA
DNA precipitation method
transformation
Escherichia coli
