RT-PCR法测定胎鼠大脑皮质内皮素-1 mRNA

RT-PCR for analysis of endothelin-1 mRNA in fetal mouse cortex

目的 建立一种检测先天性人巨细胞病毒感染胎鼠大脑皮质内皮素 1(ET 1)mRNA的方法。方法 采用逆转录 聚合酶链式反应 (RT PCR)方法并对实验条件进行优化。建立稳定的检测体系后 ,对胎鼠大脑皮质ET 1mRNA进行检测 ,并以RT PCR后产物的OD值来反映起初ET 1mR NA的模板相对含量。结果 在一定TaqDNA酶、镁离子浓度、引物浓度等条件下 ,ET 1mRNA经RT PCR扩增后产物的OD值与ET 1mRNA模板量成正比。结论 优化条件后的RT PCR是检测ET 1mRNA的一种敏感、简便、特异、可靠的方法。

Objective To establish a method to determine the pathogenic roles of endothelin-1(ET-1) mRNA in fetal mouse cortex following congenital human cytomegalovirus(HCMV) infection. Methods After the conditions used in reverse transcriptase polymerase chain reaction(RT-PCR) were optimized, ET-1 mRNA at gene transcripition level was analysed. ET-1 mRNA quantity was determined by the OD value of PCR product. Results Under the concentration of Taq DNA polymerase, the OD value of RT-PCR product was correlated with the amount of ET-1 mRNA templet. Conclusion RT-PCR is very sensitive, simple, specific and credible in determining ET-1 mRNA.