The levels of <i>SMN2</i> full-length mRNA and protein decrease during testis cell primary culture.

<p>(A) Total RNA was isolated from primary testis cells cultured for different time periods (2 hours, 48 hours, 96 hours and 24 days) and subjected to RT-PCR to amplify <i>SMN2</i> FL and Δ7 mRNAs. For comparison, total RNA isolated directly from the testis and liver was also analyzed. A representative result of three independent experiments was shown. The result showed that primary testis cells after a 2-hour culture still expressed high level of <i>SMN2</i> FL mRNA. However, the level decreased after longer cultures. (B) Proteins extracted from primary testis cells cultured for 2 hours and 96 hours were subjected to Western blotting to detect SMN protein. The result showed that SMN protein level was high in 2-hour cultured cells and decreased dramatically in 96-hour cultured cells, consistent with the result of <i>SMN2</i> exon 7 splicing. (C) snRNP complexes were isolated from primary testis cells cultured for 2 hours and 96 hours by anti-Sm antibodies. Various snRNAs were then extracted and quantitated by real-time PCR. The result showed that the levels of snRNP complexes decreased in 96-hour cultured cells compared with 2-hour cultured cells. Error bars represent standard deviation. (*) <i>P</i> < 0.05, compared with 2-hour cultured cells.