携带SKP2基因的shRNA表达质粒的构建与鉴定

Construction and appraise of SKP2 gene shRNA expression plasmid

目的构建含人SKP2基因不同位点的一系列shRNA真核表达质粒,并进行酶切鉴定、测序。方法设计合成3对SKP2基因干扰寡核苷酸序列,形成双链后将其依次连入带有U6启动子的pSIREN-RetroQ载体,构建成能产生SKP2短发卡RNA的质粒。结果构建的核苷酸序列经鉴定构建成功,能成功的克隆入带有pSIKEN RetroQ的载体中。结论成功构建并鉴定了针对人SKP2基因3个不同位点的shRNA的真核表达质粒,为SKP2为靶点的肿瘤基因治疗研究奠定了基础。

Objective To construct and appraise a series of shRNA eukaryotic expression plasmid which contains vary spots of human SKP2 gene. Methods Three pairs of hairpin-like oligonucleotide sequences specific for human SKP2 gene were designed and synthesized. They were cloned into the eukaryotic expression pSIREN-RetroQ plasmid. The recombinated plasmids were confirmed by PCR and sequencing. Results The shRNA sequences were constructed successfully, and could be inserted into the eukaryotic expression vector pSIREN-RetroQ. Conclusions Three shRNA transcripting plasmids pSIREN-RetroQ are successfully constructed, and form a basis for research in cancer gene target therapy against SKP2 gene.