克隆(编程)
生物
克隆载体
图书馆
聚合酶链反应
计算生物学
分子克隆
基因组文库
功能基因组学
基因组学
聚合酶
DNA
遗传学
计算机科学
基因组
基因
互补DNA
基序列
16S核糖体RNA
程序设计语言
作者
Jiayuan Quan,Jiajun Tian
出处
期刊:Nature Protocols
[Springer Nature]
日期:2011-02-01
卷期号:6 (2): 242-251
被引量:268
标识
DOI:10.1038/nprot.2010.181
摘要
High-throughput genomics, proteomics and synthetic biology studies require ever more efficient and economical strategies to clone complex DNA libraries or variants of biological modules. In this paper, we provide a protocol for a sequence-independent approach for cloning complex individual or combinatorial DNA libraries, and routine or high-throughput cloning of single or multiple DNA fragments. The strategy, called circular polymerase extension cloning (CPEC), is based on polymerase overlap extension and is therefore free of restriction digestion, ligation or single-stranded homologous recombination. CPEC is highly efficient, accurate and user friendly. Once the inserts and the linear vector have been prepared, the CPEC reaction can be completed in 10 min to 3 h, depending on the complexity of the gene libraries.
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