3GC cloning: PCR products cloning mediated by terminal deoxynucleotidyl transferase.
3GC cloning: PCR products cloning mediated by terminal deoxynucleotidyl transferase.
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DOI：
10.1016/j.ab.2008.03.053
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发表时间：
2008-07
影响因子：
2.9
通讯作者：
Dong Zheng;Xuedong Liu;Yanna Zhou
中科院分区：
文献类型：
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作者：
Dong Zheng;Xuedong Liu;Yanna Zhou
我们开发了一种PCR产物克隆策略，称为3GC克隆。通过这种策略，通过末端脱氧核苷酸转移酶（TdT）在3 '端含有3个以上的同聚脱氧胞苷（dCs）的PCR产物将与3G载体上两个识别位点通过SfiI偶联切割产生的3个脱氧鸟苷（dG）突出末端互补。在细菌中，结扎后插入载体连接处的多余悬垂会被修剪和修复，这种连接处的长度仅为3个G/C对。任何长度为三个或更多核苷酸的链，都可以作为TdT的底物，理论上可以通过这种策略克隆。
We have developed a PCR product cloning strategy called 3GC cloning. Through this strategy, PCR products tailed with more than three homopolymeric deoxycytidines (dCs) at the 3′ ends by terminal deoxynucleotidyl transferase (TdT) would anneal complementarily with three-deoxyguanosine (dG) protruding ends of 3G vector, which was generated through coupled SfiI cleavage at both recognition sites. Redundant overhangs at insert–vector junctions after ligation would be trimmed off and repaired in bacteria, and the length of such junction was only three G/C pairs. Any chain lengths of three or more nucleotides, which are able to serve as substrates for TdT, could theoretically be cloned by this strategy.