siRNA抑制卵巢癌细胞株SKOV3 HPA基因表达的研究  

Inhibitory effect of siRNA on HPA gene expression in SKOV3 cells

作　　者：辛晓燕[1] 毛敬[1] 刘玉[1] 于月成[1] 李萍[1] 

机构地区：[1]第四军医大学西京医院妇产科,西安710032

出　　处：《中国肿瘤生物治疗杂志》2006年第2期124-128,共5页Chinese Journal of Cancer Biotherapy

基　　金：中国博士后科学基金资助(2005037776)

摘　　要：目的：研究siRNA对卵巢癌细胞株SKOV3中乙酰肝素酶(heparanase,HPA)基因表达的抑制作用。方法：设计合成两对HPA编码基因的反向重复序列,中间间隔9个核苷酸序列,通过定向克隆至载体PGenesil-1,构建siRNA真核表达载体,经稳定转染SKOV3细胞后应用RT-PCR、real-time PCR及免疫组化技术检测卵巢癌细胞中HPA基因mRNA及蛋白表达水平的抑制情况。结果：测序证实成功地构建了编码2条shRNA的siRNA真核表达载体。通过RT-PCR、real-time PCR及免疫组化技术检测转染siRNA的SKOV3细胞,与对照组相比,HPA mRNA表达水平和蛋白表达水平明显降低。结论：构建的PGenesil-1(+)-HPA重组质粒能有效的抑制HPA基因在卵巢癌细胞株SKOV3中的表达,为研究HPA基因在肿瘤细胞中的调节途径和肿瘤的基因治疗提供了新的方法。Objective： To study the inhibitory effect of siRNA on Heparanase （HPA） expression in SKOV3 cells. Methods： Two pairs of 21 bp reverse repeated sequence targeting HPA RNA （ spaced by 9 bp nucleotide） were synthesized and were cloned into plasmid PGenesil-1 to construct recombinant plasmid PGenesil-1 （ ＋ ） -HPA expressing 2 hairpin siRNAs. The inhibition of HPA gene was detected by RT-PCR, real-time PCR and immunohistochemical staining after PGenesil-1 （ ＋ ）-HPA was stably transfected into SKOV3 cells. Results： The recombinant plasmid PGenesil-1 （ ＋ ）-HPA （expressing 2 hairpin siRNAs） was successfully constructed. RT-PCR, real-time PCR and immunohistochemical staining showed that there was a significant decrease in HPA mRNA and protein level in experimental group compared with those in control group. Conclusion： siRNA targeting HPA mRNA can specically suppress the expression of HPA gene in SKOV3 ceils; RNA interference method provides a new way for studying the role of HPA and gene therapy of cancer.

关 键 词：小分子干涉RNA HPA基因 卵巢癌 

分 类 号：R730.54[医药卫生—肿瘤]