负载双片段survivin短发夹状RNA质粒载体的构建与鉴定

目的:设计构建负载双片段survivin短发夹状RNA(shRNA)的质粒载体,为进行前列腺癌(PCa)基因治疗的研究奠定基础。方法:分别设计2条靶向survivin基因的shRNA,克隆至质粒载体中,再分别提取质粒酶切后回收、连接,构建重组质粒,提取重组质粒酶切鉴定、测序分析。结果:负载不同单片段survivinshRNA的质粒载体分别用EcoRⅠ及SacⅠ酶切鉴定,证实设计的2条单片段survivinshRNA已经分别插入目的质粒中;基因重组后,重组质粒载体用BamHⅠ酶切鉴定,证实双片段survivinshRNA均已插入重组质粒载体中;测序结果证实重组质粒载体插入的双片段survivinshRNA序列均正确无误。结论:负载双片段survivinshRNA的RNA干扰重组质粒载体构建成功。

Construction and Identification of Plasmid Vector Encoding Two Survivin ShRNA

Objective: To construct and identify the eukaryotic expression plasmids encoding two short hairpin RNA(shRNA) of survivin for the purpose of paving the way for the studies of targeted gene therapy for prostatic carcinoma(PCa).Methods: Two shRNA of survivin were designed and synthesized respectively,and then both were cloned into plasmids.Finally,the recombinant plasmids were confirmed by sequencing and agarose gel electrophoresis after restriction digestion.Results: The recombinant plasmids encoding two survivin shRNA were constructed and the aim sequence obtained.Conclusion: Successful construction of the recombinant provides a sound basis for the research of targeted gene therapy for PCa.Natl J Androl,2006,12(6):512-515