Enhancement of the reporter gene repression by a Flag-Piwil2 transgene. (A) Demonstration of increased PIWIL2-associated piRNAs in Flag-Piwil2 BAC transgenic testes at 5 wk. PIWIL2- and PIWIL1-bound piRNAs are indicated. Northern blotting was performed with a probe detecting a pachytene piRNA from the Watson cluster. 5S RNA and ethidium bromide staining are the loading controls. (B) qRT-PCR of beta-geo [Gt(ROSA)26Sor] and EGFP mRNAs. Germ cells were isolated from Gt(ROSA)26Sor and Cag-EGFP mice with or without EGFP-neo and also with or without Flag-Piwil2. A significant difference in beta-geo expression is observed between EGFP-neo/+ germ cells with or without Flag-Piwil2. The level of beta-actin mRNA was used as a reference. (*) P < 0.05, Student's t-test. Error bars indicate the standard deviations (SDs) of biological replicates.
