40 cycles of 97°C for 30 seconds followed by 59.7°C for 1 min. To measure the quantity of a given RNA species, the threshold cycles (Ct) were monitored by the Applied Biosystems 7900HT Fast Real-Time PCR System. Each mRNA expression was calculated by relative quantification using a normalization factor (geometric mean of two reference genes selected by geNORM program, http://medgen.ugent.be/genorm/) [35] and qBASE program for internal and external calibration and also for easy care of large RT-PCR datasets (http://medgen.ugent.be/qbase/). According to our analysis of reference genes [36] the beta-actin (ACTB) and ribosomal large P0 (RPLP0) genes for PFC, and the 18S and RPLP0 genes for the MC were chosen for normalization.
