Figure 1

Aromatase expression in mouse fetal testis. A. Scheme showing the different testis aromatase variants (T1, T2 and T3). Aromatase exons from exon 2 to exon 10 are represented (gray boxes) for each transcript variant T1, T2 and T3. For each variant the relative position of the RT-PCR primers (red arrows), quantitative qRT-PCR (black arrows) and internal probes (blacks lines) used are represented. B. Expression of different aromatase transcript variants (T1, T2 and T3) during mouse testis development. Products were generated by RT-PCR with the primer combinations E2-E9/10 (see Table 1) and visualized on 2% agarose gel. NC: 3 negative control (PCR master mix without template). M: DNA ladder; dpc: days post-coitum; dpp: days post-partum. C. Absolute quantitative expression of aromatase transcripts during mouse testis development. Total mRNA was isolated from fetal testes at the indicated stages of development, reverse transcribed and aromatase expression was quantified by real-time PCR using the TaqMan method. This method allows the identification of each isoform by using a specific internal probe (see Figure 1A for position and Table 1 for sequences). In order to calculate precisely the copy number of each transcript, each transcript was isolated on gel, re-amplified and quantified. For each experiment a standard curve was constructed using the isolated transcript as template with their corresponding qRT-PCR primers. Data shown are the mean ± SEM (n = 4-6). ^*P < 0.05 (Student’s t-test; compared to the mRNA copy number at 13.5 dpc). D. Aromatase protein expression during mouse testis development. On western blots, three protein isoforms were recognized by the anti-aromatase antibody (MCA2077T, Serotec, France). Lane A, ovary from a pregnant mouse: only one band that corresponds to the aromatase full-length protein. Lane B, 2 dpp mouse testis; lane C and D, extracts from 15.5 and 13.5 dpc mouse testes, respectively.