title: Generation of a germ cell-specific mouse transgenic CHERRY reporter, Sohlh1-mCherryFlag.
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.
authors.affiliation: Magee-Womens Research Institute, Department of Obstetrics, Gynecology, and Reproductive Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania 15213, USA.
authors.forename: Hitomi
authors.initials: H
authors.lastname: Suzuki
authors.forename: Christina Tenenhaus
authors.initials: CT
authors.lastname: Dann
authors.forename: Aleksandar
authors.initials: A
authors.lastname: Rajkovic
mesh_terms: D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics*; D022202:Chromosomes, Artificial, Bacterial / Q000235:genetics; D005260:Female; D005456:Fluorescent Dyes; D018390:Gene Targeting; D017930:Genes, Reporter / Q000235:genetics; D005854:Germ Cells / Q000378:metabolism*; D008164:Luminescent Proteins / Q000235:genetics*; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D000097573:Red Fluorescent Protein
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D005456:Fluorescent Dyes; D008164:Luminescent Proteins; D000097573:Red Fluorescent Protein; C511466:Sohlh1 protein, mouse