A novel series of high-efficiency vectors for TA cloning and bluntend cloning of PCR products
A novel series of high-efficiency vectors for TA cloning and bluntend cloning of PCR products
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DOI：
10.1038/s41598-019-42868-6
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发表时间：
2019-04-23
影响因子：
4.6
通讯作者：
Motohashi, Ken
中科院分区：
文献类型：
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作者：
Motohashi, Ken
高效的PCR克隆方法是现代分子生物学中不可缺少的，因为它可以大大提高DNA克隆过程的效率。在这里，我描述了TA克隆和平末端克隆的三种载体的发展。具体地，设计pCRT和pCRZeroT以提高TA克隆的效率。pCRZeroT也可以与pCRZero一起使用，以促进使用ccdB基因的平末端克隆。使用pCRZero和pCRZeroT，并应用Golden Gate反应，我开发了一种直接PCR克隆方案，使用未消化的环状载体和PCR产物。这种直接PCR克隆方案产生的集落形成率和克隆效率与用预消化载体和PCR产物通过常规PCR克隆获得的集落形成率和克隆效率相当。我设计的三种质粒可从Addgene（https：//www.addgene.org/）获得。
An efficient PCR cloning method is indispensable in modern molecular biology, as it can greatly improve the efficiency of DNA cloning processes. Here, I describe the development of three vectors for TA cloning and blunt-end cloning. Specifically, pCRT and pCRZeroT were designed to improve the efficiency of TA cloning. pCRZeroT can also be used with pCRZero to facilitate blunt-end cloning using the ccdB gene. Using pCRZero and pCRZeroT and applying the Golden Gate reaction, I developed a direct PCR cloning protocol with non-digested circular vectors and PCR products. This direct PCR cloning protocol yielded colony-formation rates and cloning efficiencies that are comparable with those obtained by conventional PCR cloning with pre-digested vectors and PCR products. The three plasmids I designed are available from Addgene (https://www.addgene.org/).