Bulk RNA-seq of fetal and adult murine Leydig cells

To compare expression profiles of fetal and adult mouse Leydig cells, we performed bulk RNA sequencing. Mouse testis from XY Nr5a1 Tg mouse embryos at embryonic day (E) 18.5 and 100-day old mice were harvested. Tissues were enzymatically dissociated and then filtered.The single cell solution is then sorted by fluorescent-active cell sorting to obtain a solution of alive and single Nr5a1-GFP+ cells. RNA was extracted from these cells with RNeasy Mini kit (Qiagen). Sequencing libraries were prepared with the TruSeq Stranded Total RNA Library Prep Gold (Ribo-Zero) and sequenced on an Illumina HiSeq 2500 (50 bp, paired end, ~35 million reads expected). Each condition was performed in three independent replicates.