Fig. 2
Ranking of proposed and commonly (previously) used reference genes in placental, luteal and healthy uterine samples, generated by RefFinder (a) and NormFinder (b) applications. The NormFinder tool was used to calculate intragroup (group refers to a particular tissue: 1-CL, 2-placenta, 3-uterus) stability values. Both softwares ranked KDM4A, EIF4H and PTK2 as the most stably expressed genes. The comparison was made including previously used reference genes (GAPDH, β-actin, cyclophilin A/PPIA) (c) Determination of the optimal number of control genes for normalization in canine CL, placenta and uterus. Pairwise variation (Vn/n + 1) analysis was done to determine the number of control genes required for accurate normalization. A cut-off value 0.15 was applied (Vandesompele et al. 2002). Pairwise variation analysis shows that V2/3 values in canine CL and placenta, and V3/4 in the uterus, were lower than 0.15, indicating that two reference genes are suitable for gene normalization in CL and placenta, but 3 genes should be included for uterus