摘要
目的 构建CD14 7反义RNA表达质粒载体。方法 用DNA重组技术将人CD14 7基因反向克隆到真核表达质粒PCD NA3.1中 ,构建成CD14 7反义RNA表达质粒PCDNAasCD14 7。用阳离子脂质体介导转染人卵巢癌细胞系 8910 ,经G4 18筛选后获得的克隆进行鉴定。结果 CD14 7反义RNA表达质粒载体PCDNAasCD14 7经限制性酶切及部分序列分析证明基因插入正确。流式细胞仪及免疫组化SP法染色均证实 :转染细胞 8910 /PCDNAasCD14 7,与亲本细胞相比 ,CD14 7的表达明显下降。结论 成功构建了CD14 7反义RNA表达质粒载体PCDNAasCD14 7,此实验结果为进一步研究CD14 7蛋白分子的生物学功能以及为卵巢癌的基因治疗研究奠定了基础。
Objectives To construct eukaryotic antisense RNA expression vector of CD147 and to identify the clones. Methods PCDNAasCD147 was constructed by inserting CD147 cDNA reversely to eukaryotic expression vector PCDNA3.1 .The ovarian cancer cell strain 8910 was transfected by PCDNA as CD147. After selected by G418,positive clones were identified by fluorocytometry assay and indirect immunofluorescence staining. Result It was verified by partial nucleotide sequencing and restriction endonuclease digestion that the constructed eukaryotic antisense RNA expression vector PCDNAasCD147 was correct. Indirect immunofluorescence staining and fluorocytometric assay showed that the expression of CD147 was significantly suppressed in 8910 cells transfected by PCDNAasCD147 as compared with the untransfected 8910 cells. Conclusion The results of this study lay the foundation for further studying the biological functions of CD147 and gene therapy of ovarian cancer.
出处
《肿瘤》
CAS
CSCD
北大核心
2002年第3期194-196,F003,共4页
Tumor
