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目的制备荧光定量聚合酶链反应(PCR)方法进行人急性早幼粒细胞白血病(APL)融合基因PMLRARα(bcr1型)mRNA表达量分析的RNA标准品,为白血病微小残留状态(MRD)的监测奠定基础。方法体外转录RNA的方法制备目的基因PMLRARα及管家基因ABL(Ablesongene)RNA标准品。结果目的基因PMLRARα的RNA标准品浓度分别为1×106、1×105、1×104、1×103、1×102拷贝/μl时CT(thresholdcycle,循环阈值,即产生可被检测到荧光信号所需的最少循环数)值分别为20.09、23.58、26.35、29.63、33.48。标准曲线的斜率为-3.30,相关系数为0.999。管家基因ABL标准品浓度分别为1×106、1×105、1×104、1×103、1×102拷贝/μl时,CT值分别为17.27、20.49、24.00、27.28、30.41。标准曲线的斜率为-3.24,相关系数为1.000。结论构建的两个RNA标准品可以得到良好的标准曲线,且标准曲线显示线形关系良好,标准品构建成功。
Objective To prepare an RNA standard for the quantitative analysis of PMLRARα (bcr1) mRNA expression in human acute promyelocytic leukemia (APL) by fluorescence quantitative polymerase chain reaction (PCR) and to lay the foundation for the monitoring of minimal residual disease (MRD) in leukemia basis. Methods The target gene PMLRARα and ABL (Ablesongene) RNA standard were prepared by in vitro transcription of RNA. Results The target RNA of PMLRARα was 1 × 106, 1 × 105, 1 × 104, 1 × 103 and 1 × 102 copies / μl, respectively. CT (threshold cycle) The minimum number of cycles required) values were 20.09,23.58,26.35,29.63,33.48. The slope of the standard curve is -3.30 and the correlation coefficient is 0.999. The concentrations of housekeeping gene ABL were 1 × 106, 1 × 105, 1 × 104, 1 × 103 and 1 × 102 copies / μl, respectively. The CT values were 17.27,20.49,24.00,27.28,30.41 respectively. The slope of the standard curve is -3.24 and the correlation coefficient is 1.000. Conclusion The constructed two RNA standards can get a good standard curve, and the standard curve shows a good linear relationship, the standard was successfully constructed.