GSM2157630: rep1_GES14_04762_ATCACG; Mus musculus; RNA-Seq
Each enriched population (4 biological replicates) of pachytene/diplotene spermatocytes was obtained from the testes of 6 9-week old mice by sedimentation at unit gravity [3] (Bellvé, 1993). Mixed germ-cell suspensions were prepared as described above, and after the three 0.5% BSA/KRB washes, cells were allowed to separate by cellular sedimentation at unit gravity in a 2–4% BSA gradient generated over 2.5 h in a STA-PUT apparatus (ProScience Inc., GlassShop, Toronto, ON, Canada).  Following the sedimentation process, 10 ml fractions were collected and examined using light microscopy and differential interference contrast Nomarski optics. Cells were identified based on morphological criteria and size [3] (Bellvé 1993). Populations of pachytene/diplotene spermatocytes of average purity= ~90% were collected. For every cell separation, and for each population of cells collected, an aliquot of cells was snap-frozen for subsequent RNA extraction. Isolated germ cell samples were resuspended in QIAzol Lysis Reagent according to the manufacturer's instructions, and total RNAs were purified from homogenized cells using Qiagen RNeasy Mini Kit (74104).