实时荧光定量反转录聚合酶链反应检测bcl-2基因mRNA表达

Real-time quantitative RT-PCR for the detection of bcl-2 mRNA expression

Objective To establish a real time quantitative RT PCR method for the detection of expression of bcl 2 mRNA. Methods The vector containing bcl 2 gene as standard template was constructed with T A cloning technique. The fluorogenic probe (i.e.,Taq man probe) was used to establish a real time RT PCR. bcl 2 mRNA expression level in Burkitt's lymphoma pre and post treated with bcl 2 antisense phosphothioate oligodeoxynucleotides (AS PS ODN) was determined with real time quantitative RT PCR,also with semi quantitative RT PCR. Results The expression of bcl 2 mRNA in Burkitt's lymphoma treated with bcl 2 AS PS ODN decreased significantly, and no changes of bcl 2 mRNA expression in group treated with nonsense ODN were noticed. The semi quantitative RT PCR results also demonstrated that bcl 2 level varied as detected with real time fluorogenic quantitative RT PCR, but less sensitive and accurate. Conclusion Detection of bcl 2 mRNA expression with the fluorogenic real time quantitative RT PCR is more accurate and sensitive than semi quantitive RT PCR.