Validation of endogenous control genes in lasered and unlasered mice eyes.

<p>5–8-week-old female C57BL/6 mice were prepared for validation of such endogenous control genes as m-cyclophilin A, beta-actin and gapdh. Three days after photocoagulation at 10 locations per eye, the mRNA expression levels of these three housekeeping genes in eyecups were compared with those of age-matched unlasered eyes. In order to determine whether the expression of the potential internal control gene varies under the experimental conditions, the realtime PCR data was presented from the replicate cDNAs as 2^−Ct and Student's t-tests were performed between the treated and untreated groups as described <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0094743#pone.0094743-Schlingemann1" target="_blank">[9]</a>. The fold changes in the three genes in the treated samples compared to the untreated samples were 1.44 for β-actin, 0.94 for cyclophilin A, and 0.06 for gapdh. Student's t-tests showed that there was significant difference between the treated and untreated samples in gapdh expression (P<0.01), but none in β-actin (P = 0.14) or cyclophilin A (P = 0.93). This suggests that cyclophilin A is presumably the most suitable endogenous control among these three genes.