中文摘要：

目的：构建人端粒酶逆转录酶（hTERT）启动子调控的趋化因子受体4（CXCR4）的RNA干扰逆转录病毒载体，并研究其在hTERT^＋前列腺癌细胞、乳腺癌细胞中的表达。方法：用PCR扩增CXCR4特异性小干扰RNA（small interfering RNA，siRNA）．转入带有增强型绿色荧光蛋白（EGFP）和启动子U6的Pgensil-1质粒。以hTERT启动子代替U6启动子．再将重组基因片段导入到逆转录病毒真核表达载体PLXSN中,行限制性酶切鉴定。转染PA317病毒包装细胞．收获病毒上清分别转染前列腺癌细胞PC-3m、LNCaP及人胚肺成纤维细胞MRC5和人乳腺癌细胞MCF7．RT—PCR检测CXCR4mRNA表达。结果：限制性酶切鉴定该重组质粒，成功构建了PLXSN／EGFP—hTERT—siCXCR4。与空载体组比较．PC-3m、LNCaP、MCF7细胞中CXCR4-siRNA对CXCR4 mRNA的48h抑制率分别为（87．02±10．34）％、（61．70±9．77）％、（88．31±9．20）％，、MRC5细胞中PEXSN—hTERT—siCXCR4对CXCR4mRNA不抑制。结论：该逆转录病毒系统中hTERT启动子调控的下游RNA干扰序列可选择性地在hTERT＋前列腺癌细胞、乳腺癌细胞中表达，在hTERT-MRC5细胞不表达．具有显著的靶向性。

英文摘要：

Objective： To construct RNA interfering retrovirus veetor targeting CXCR4 gene driven by human telomerase reserve transeriptase gene promoter and investigate CXCR4 expression in hTERT＋ carcinoma cells. Methods： The CXCR4 targeting siRNA gene was cloned by PCR. The PCR products were inserted into Pgensil-1 plasmid containing U6 promoter and EGFP. U6 promoter was replaced by hTERT promoter. Then, the recombinant EGFP-hTERT-siCXCR4 fragment was sub-cloned into PLXSN and evaluated by restriction enzyme. The virus obtained from transfected PA317 cells was transfected respectively into PC-3m,LNCaP,MCF7,MRC5 cells. The expression of CXCR4 mRNA was detected by RT-PCR. Results：The expression of CXCR4 mRNA in PC-3m,LNCaP and MCF7 were reduced by the recombinant PLXSN-hTERT-siCXCR4 constructed successfully. The inhibitory rates of the expression of CXCR4 mRNA were（87.20±10.34）%, （61.70 ± 9.77）%, （88.31±9.20）% at 48th hour respectively.CXCR4 mRNA was not inhibited by PLXSN-hTERT-siCXCR4 in MRC5 cells. Conclusion： The downstream CXCR4-siRNA controlled by hTERT promoter in retrovirus system could be expressed selectively in hTERT＋ carcinoma cells, but not in hTERT-cells, which shows obvious targeting characters.