Pretheeban, T., Gordon, M. B., Singh, R. and Rajamahendran, R. 2011. Comparison of expression levels of candidate genes in endometrium of dairy heifers and lactating dairy cows. Can. J. Anim. Sci. 91: 255-264. Pregnancy rates (PR) in high-producing lactating dairy cows have declined drastically over the past several decades, but those of heifers have remained constant. Reduced PR could be due to multiple causes, and the underlying pathophysiological mechanisms are still unclear. A compromised maternal uterine environment could be one of factors that could affect the PR. This study was performed to compare the nature of the uterine environment in dairy heifers and lactating dairy cows (2nd/3rd parity) by analyzing the expression levels of selected endometrial genes. Estrus was synchronized in heifers (n=5) and lactating dairy cows (n=5) and endometrial biopsies were performed during the mid luteal phase (day 11) of the estrous cycle. Real-time polymerase chain reaction (Q-RT PCR) and immunohistochemistry were performed to analyse the mRNA and protein levels of genes respectively. Relative abundance of BCL2, HSPA1A, IL1A, TNF, IGF1, FGF2 and SERPINA14 transcripts and the protein expression of IL1A, TNF and FGF2 were significantly higher in heifers in comparison with lactating dairy cows. Our findings suggest an altered endometrial environment in lactating dairy cows compared with heifers. However, whether these differences play a role in pregnancy outcomes should be further investigated.