ZNF580基因原核表达载体的构建与鉴定

【目的】构建ZNF580基因原核表达载体,为获得大量重组ZNF580蛋白并制备其多克隆抗体奠定基础。【方法】根据ZNF580cDNA序列的开读框架设计上、下游引物,利用PCR技术扩增ZNF580开放阅读框cD-NA序列,将PCR扩增的目的片段与载体pET30a分别双酶切、回收纯化后做定向连接,EcoRI、XhoI双酶切电泳筛选、鉴定并测序。【结果】双酶切pET30a-ZNF580,电泳分析插入片段长度为524 bp,与预期的结果一致,经连接点两端进行测序,测序结果显示接头两端序列连接正确。【结论】成功构建原核表达载体pET30a-ZNF580。

Construction and identification of prokaryotic expression vector pET30a-ZNF580

[Objective] To construct a prokaryotic expression vector pET30a-ZNF580. [Methods] The primers were designed according to the open reading frame of ZNF580 gene cDNA sequence, the digestion of the PCR fragment and pET30a with both EcoR I and XhoI was hgated. Recombinant prokaryotic expression vector pET30a-ZNF580 was identi- fied by restriction enzymes digestion, agarose gel electrophoresis and DNA sequencing. [Results] Restriction enzymes di- gesting and DNA sequencing showed that the sequence of the recombinant plasmid pET30a-ZNF580 was correct. [Conclusions] The recombinant prokaryotic expression vector pET30a-ZNF580 has been successfully constructed.