【摘要】 背景与目的:全基因组相关联研究发现了染色体10q11区域的单核苷酸多态性(single nucleotide polymorphism,SNP)位点rs10993994与前列腺癌的遗传易感性相关,本研究探讨rs10993994影响前列腺癌发病风险的可能机制,即研究rs10993994对前列腺癌细胞微精液蛋白β(microseminoprotein beta,MSMB)基因转录活性的影响。方法:化学合成MSMB基因的启动子序列,由于位于序列中间的SNP位点rs10993994有2种等位基因(T/C)可能,合成2条启动子MSMB promoter-T和MSMB promoter-C。将2条启动子通过酶切的方法导入荧光素酶报告基因载体(pGL3-basic vectors)中,筛选阳性克隆后,将携带启动子的质粒转染入前列腺癌细胞PC-3和LNCaP中,最后通过荧光检测仪检测重组质粒中启动子的转录活性。结果:成功合成2条启动子序列:MSMB promoter-T和MSMB promoter-C;分别将启动子序列与荧光素酶报告基因载体重组,形成重组质粒pGL3-MSMB promoter-T和pGL3-MSMB promoter-C。在前列腺癌细胞PC-3中,重组质粒MSMB promoter-C组的相对荧光度为2.27±0.39,显著高于重组质粒MSMB promoter-T组(0.57±0.13),差异有统计学意义(P<0.05);在前列腺癌细胞LNCaP中,重组质粒MSMB promoter-C组的相对荧光度为1.70±0.32,显著高于重组质粒MSMB promoter-T组(0.37±0.09),差异有统计学意义(P<0.05)。结论:重组质粒pGL3-MSMB promoter-C的转录活性强于重组质粒pGL3-MSMB promoter-T的转录活性;rs10993994可以影响到MSMB基因启动子的启动转录活性,rs10993994位点上等位基因胞嘧啶(C)较胸腺嘧啶(T)更能促使MSMB基因转录。更多还原

【Abstract】 Background and purpose: Rs10993994 at 10q11 was found to be associated with prostate cancer risk by genome-wide association studies. This study tried to reveal its mechanism and to explore the role of rs10993994 for the transcriptional activity of microseminoprotein beta(MSMB) gene in prostate cancer cell lines.Methods: Promoter fragments were generated by chemical synthesis. Due to the two possibility of rs10993994(T/C) in the region, we generated two promoter fragments: MSMB promoter-T and MSMB promoter-C. The fragments were then cloned into pGL3-basic vectors, positive clones were transfected into prostate cancer cell lines PC-3 and LNCaP, finally, relative level of fluorescence was detected by fluoresce detector. Results: We generated two promoter fragments of MSMB, MSMB promoter-T and MSMB promoter-C. The two promoter fragments were cloned with pGL3-basic vectors to pGL3-MSMB promoter-T and pGL3-MSMB promoter-C. In PC-3, the relative level of fluorescence of pGL3-MSMB promoter-C was significant higher than that of pGL3-MSMB promoter-T(2.27±0.39 vs 0.57±0.13, P<0.05); In LNCaP, the relative level of fluorescence of pGL3-MSMB promoter-C was significant higher than that of pGL3-MSMB promoter-T(1.70±0.32 vs 0.37±0.09, P<0.05). Conclusion: The transcriptional activity of pGL3-MSMB promoter-C was stronger than that of pGL3-MSMB promoter-T. Rs10993994 could influence the transcriptional activity of MSMB gene promoter in prostate cancer cell lines PC-3 LNCaP, allele C in rs10993994 could facilitate transcription than T.更多还原