Abstract
Visualization of differentiating germ cells is critical to understanding the formation of primordial follicles in the ovary, and the commitment of spermatogonial stem cells to differentiation. We engineered and generated a BAC transgenic mouse line, Sohlh1-mCherryFlag (S1CF), under the direction of the native Sohlh1 promoter. Sohlh1 is a germ cell-specific gene that encodes the basic helix-loop-helix (bHLH) transcriptional regulator that is essential in oogenesis and spermatogenesis. Sohlh1 expression is unique, and is limited to perinatal and early follicle oocytes and differentiating spermatogonia. The Sohlh1-mCherryFlag transgene was engineered to fuse SOHLH1 to the red fluorescent protein CHERRY with 3-tandem-FLAG tags. S1CF animals fluoresce specifically in the oocytes of perinatal ovaries and small follicles in adult ovaries, as well as in spermatogonia, a pattern that is similar to endogenous SOHLH1. Moreover, S1CF rescued germ cell loss and infertility in both male and female Sohlh1(-/-) animals. The FLAG-tag on S1CF was effective for immunostaining and immunoprecipitation. The Sohlh1-mCherryFlag transgenic mouse provides a unique model to study early germ cell differentiation, as well as in vivo imaging and purification of differentiating germ cells.