Establishment of a real-time fluorescence quantitative RT-PCR for detecting WT1 mRNA in leukemia cells

Objective To establish a real-time fluorescence quantitative RT-PCR(RQ-PCR) method for detecting the expression of WT1 gene in leukemia cells. Methods A specific fluorescence-labeled probe capable of binding to cDNA target was i ncorporated into the reaction system of a regular RT-PCR. The RQ-PCR method fo r WT1 mRNA was set up and the specificity and sensitivity were tested. Res ults The specificity and sensitivity of this RQ-PCR were ideal. Sensit ive experiments revealed that 10 -4 WT1 mRNA could be detected. The CV of t his method was 1 93%. The standard curve indicated that there was a linear rela tionship between CT value and the logarithmic value of template concentration (s lope=-3 5, r=0 997). With this method the accurate quantitative assessment of initial amount of WT1 mRNA was performed. Conclusion RQ-PC R is proved to be a sensitive, specific, reliable and convenient method for the detection of WT1 mRNA in leukemia cells.