荧光定量PCR检测前列腺特异膜抗原及其变异体DNA的方法学

Methodology of Fluorescent Quantitative PCR to Test PSMA and Alternative Spliced Variant PSMA5 DNA

目的拟建立以荧光定量逆转录聚合酶链反应(FQ-RT-PCR)检测前列腺特异性膜抗原(PSMA)及其变异体5(PSMA5)DNA的定量方法。方法自行设计三对引物,自制PC3.0-T-PSMA及PC3.0-T-PSMA5质粒作为标准品,采用FQ-RT-PCR检测前列腺癌LNCap细胞中PSMA及PSMA5 DNA的含量。结果标准曲线的斜率为-0.29,相关系数为0.995,成功地扩增出前列腺癌LNCap细胞中PSMA及PSMA5基因的目标DNA片段。结论应用自行设计的引物及标准品的FQ-RT-PCR能对前列腺癌细胞株中PSMA及变异体PSMA5的DNA进行定量PCR检测。

Objective To build a FQ-PCR method which could be used to test the PSMA and the variant PSMA5 DNA.Methods Three pairs primers were designed and PC3.0-T-PSMA,PC3.0-T-PSMA5 plasmids were made as the standards,the fluorescent quantitative PCR method was used to test the PSMA and PSMA5 DNA in prostate cancer LNcap cells.Results The slope of the standard curve was 0.29 and correlation coefficient was 0.995,the target DNA fragments were amplified successfully.Conclusion PSMA and PSMA5 DNA in prostate cancer LNcap cell can be tested by FQ-RT-PCR with the primers and standards we designed.