基于特异性引物高效构建TALE文库  

Efficient construction of TALE library based on specific primers

作　　者：蔡思杰 张书衍 王晓微 郭显阳 詹硕 CAI Sijie;ZHANG Shuyan;WANG Xiaowei;GUO Xianyang;ZHAN Shuo(College of Life Science and Bioengineering,Shenyang University,Shenyang 110044,China;Shenyang Shuangding Pharmaceutical Co.Ltd.,Shenyang 110179,China)

机构地区：[1]沈阳大学生命科学与工程学院,辽宁沈阳110044 [2]沈阳双鼎制药有限公司,辽宁沈阳110179

出　　处：《高校化学工程学报》2023年第6期943-950,共8页Journal of Chemical Engineering of Chinese Universities

基　　金：国家级大学生创新创业训练计划平台-创新训练项目(202211035010X)。

摘　　要：人工构建的转录激活因子样效应物(TALE)已广泛应用于基因转录调控和基因组编辑领域,针对TALE构建过程非常复杂,研发了一种构建TALE文库的新方法。首先,基于金门酶切连接方法,设计3种用于识别碱基G的TALE骨架质粒;其次,使用4种特异性引物扩增3种骨架质粒得到质粒突变体;最后,使用金门酶切连接方法连接3种质粒突变体,成功构建最终的TALE文库。基因测序结果表明该文库包含所有可能确定TALE-DNA结合特异性的核苷酸组。由于TALE的高靶向性和多功能性,该TALE文库将广泛用于生命科学领域。Artificially constructed transcriptional activator like effectors(TALE)are widely used in gene transcription regulation and genome editing.A new method for constructing a TALE library was invented due to the complexity of TALE construction processes.Based on the golden gate restriction enzyme digestion method,three backbone plasmids of TALE were first designed to recognize the guanine(G).Three backbone plasmids were then amplified with four specific primers to obtain plasmid mutants.The final TALE library was successfully constructed by connecting three plasmid mutants with the golden gate restriction enzyme digestion method.The results of gene sequencing show the library contains all nucleotides that could determine the specificity of TALE-DNA binding.This provides the best material for researchers to use TALE directly.The TALE library can be widely used in life sciences for its high targeting and versatility.

关 键 词：TALE文库 基因转录调控 基因组编辑 特异性引物 金门酶切连接方法 

分 类 号：Q812[生物学—生物工程]