GSM6260708: Somatic cell, primary, rep50; Mus musculus; RNA-Seq
Follicles were incubated in Accutase solution (Stemcell Technologies, Canada) (Zhang et al., 2018) until somatic cells partially detached from each other. Specifically, primordial and transitioning follicles were incubated in Accutase solution for 5 minutes, and primary and secondary follicles were incubated in Accutase solution for 10 minutes on 37 °C heated stage following agitation to dissociate somatic cells from the oocyte. Primordial and transitioning follicles were pipetted with 20 μm Pasteur pipet prepared by a micropipette puller (Sutter Instrument, CA, USA), primary follicle with a 30 μm Pasteur pipet and secondary follicle with a 50 μm stripper tip (Origio Inc.). Single cells were washed in three drops of 50 μL L-15/PVA with 100 μL/mL FBS. scRNA-seq libraries were generated using SMART-Seq2 protocol (Trombetta et al., 2014). Briefly, cDNA was reversed transcribed from single cells using Maxima RT (Thermo Fisher Scientific) and whole transcriptome amplification (WTA) was performed. WTA products were purified using the Agencourt AMPure XP beads (Beckman Coulter, IN, USA) and used to prepare paired-end libraries with Nextera XT  (Illumina, CA, USA).