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Expression analysis of cyclin D2 as a potential Elf5 target gene. A. The expression pattern of cyclin D2 and Elf5 during different stages of mammary gland development. The expression profile was generated by analysis of microarray data from Stein et al. B. Relative expression of cyclin D2 mRNA by real time RT-PCR during mouse mammary gland development. Total RNA from wild type (WT) and K14-Cre/Elf5f/f (null) mouse mammary glands at pregnancy day 17.5 and lactation day 1 were analyzed for relative expression of Ccnd2 mRNA levels by real time RT-PCR. The housekeeping gene Gapdh was used to normalize gene expression. Results are from two or more independent experiments. C. Expression of cyclin D2 protein during mouse mammary gland development. Protein extract from wild type (WT) and K14-Cre/Elf5f/f knockout (KO) mouse mammary glands at pregnancy day 17.5 and lactation day 1 were analyzed by western blot with antibodies against cyclin D2 and p27. Equal loading was assessed by anti-β-tubulin antibodies and the absence of Elf5 in KO was confirmed by anti-Elf5 antibodies.