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目的建立胃肠癌中C-myc基因的荧光定量PCR(FQ-PCR)方法。方法胃肠癌细胞中的C-myc基因与质粒PGEM-T Easy Vector重组,转化大肠杆菌E.coli DH5α,获得克隆的C-myc基因标准模板。用ABI PRISM 7700PCR仪检测FQ-PCR扩增产物制成标准曲线来检测未知标本中C-myc的含量。结果 FQ-PCR扩增产物呈“S”形动力学曲线;ct(循环阈值)与PCR体系中起始模板拷贝数的对数值之间存在严格的线性关系,显示了FQ-PCR定量的准确性。结论 FQ-PCR是一种快速、简便、灵敏、准确的定量C-myc基因的方法。
Objective To establish a fluorescence quantitative PCR (FQ-PCR) method for C-myc gene in gastrointestinal cancer. Methods The C-myc gene in gastrointestinal cancer cells was recombined with plasmid PGEM-T Easy Vector and transformed into E. coli DH5α to obtain the standard template of cloned C-myc gene. The ABI PRISM 7700 PCR instrument was used to detect FQ-PCR amplification products to make a standard curve to detect the content of C-myc in unknown samples. Results FQ-PCR products showed a “S” shape kinetic curve; there was a strict linear relationship between the ct (cycle threshold) and the logarithm of the initial template copy number in the PCR system, indicating that FQ-PCR quantification accuracy. Conclusion FQ-PCR is a rapid, simple, sensitive and accurate method for quantifying C-myc gene.