Endogenous Control Genes in Prostate Cells: Evaluation of Gene Expression Using ‘Real-Time’ Quantitative Polymerase Chain Reaction
Objective: Our aims were to measure the level of expression of Abelson (ABL1), β-glucuronidase (GUS) and glucose-6-phosphate dehydrogenase (G6PD) genes in exfoliated urine cells from healthy and transrectal ultrasound biopsy patients with elevated prostate-specific antigen levels and/or abnormal digital rectal examinations or urinary symptoms indicative of prostate problems, as well as in archived formalin-fixed paraffin-embedded prostate materials. Materialsand Methods: Real-time quantitative polymerase chain reaction (RQ-PCR) was used to evaluate the suitability of the 3 control genes, i.e. ABL1, GUS and G6PD, as control genes for prostate cancer cells. Exfoliated urine cells from 30 healthy males, 53 male patients, 138 cases of archived paraffin-embedded prostate tissues and 3 prostate cell lines were sampled. All cells were lysed in guanidine isothiocyanate buffer from which RNA was extracted and converted to cDNA by random hexamer priming. RQ-PCR was performed using TaqMan chemistries. Results: There was no significant difference in the level of expression for each of the 3 control genes in the cell lines. There was a significant difference in GUS transcript level between patients and healthy controls in both urine and prostate tissue sections (p < 0.05). G6PD transcript numbers also differed significantly from those of GUS in the prostate cell lines and tissue sections (p < 0.05). The transcript numbers of all the control genes were significantly reduced in aged samples (p < 0.001). Conclusion: The ABL1 gene was the most stable control gene in both clinical specimens and cell lines. Therefore, we recommend its use to enable standardization and interlaboratory comparisons for the RQ-PCR of prostatic tumour markers.