目的设计并筛选能高效抑制人角质形成细胞膜上蛋白酶激活受体-2(PAR-2)基因表达的小干扰RNA(siRNA)序列。方法根据人PAR-2 mRNA的序列,设计合成3对针对靶基因的不同siRNA。应用脂质体Lipofectamin2000转染试剂,将3对siRNA转染人角质细胞,半定量RT-PCR技术检测PAR-2mRNA表达水平、Western blot检测PAR-2蛋白表达,以评价干扰效果。结果设计合成的3对PAR-2 siRNA中有2对siRNA可有效抑制PAR-2基因表达,以第2对siRNA效果为佳。结论成功设计合成特异且高效阻断PAR-2表达的siRNA,为进一步应用RNAi技术沉默PAR-2基因奠定实验基础。

Objective To design and screen small interfering RNA(siRNA) targeted protease-activated receptor-2(PAR-2) mRNA in Keratinocyte that can specifically and effectively suppress PAR-2 expression.Methods Three pairs of PAR-2 specific double stranded siRNAs were designed according to the sequence of PAR-2 mRNA.These siRNA were transfected into human keratinocyte by Lipofectamine 2000.The transcription level of PAR-2 was analyzed by semi-quantitative RT-PCR and PAR-2 protein expression was tested by western blot in order to evaluate the effect of RNA interference.Results Two of the three PAR-2 siRNA effectively inhibited the expression of PAR-2 and the second one is more effective.Conclusion PAR-2 siRNAs can be successfully designed and synthesized,which can specifically and effectively suppresses the expression of PAR-2 gene.The results in this study lay the foundation for further study on PAR-2 gene silencing by RNA interfere(RNAi).