GSM8835768: granulosa cell from hierarchical follicles; Gallus gallus; RNA-Seq
The GC layers of hierarchical follicles (group HF-GCs) and prehierarchical follicles (group PHF-GCs) were collected.GC layers were cut into pieces and then digested with 1 mg/ml of collagenase type IV (Sigma) in PBS for 10 min at 37 °C until cells were completely separated. Dissociated cells were ﬁltered through a 40-µm cell strainer, and were washed twice by resuspension in DMEM. Cell viability of GCs exceeded 90% after their isolation. The cell density was adjusted to the concentration of 700–1200 cells/μL. High quality single cell suspensions were subjected to 10x scRNA-seq library preparation using Chromium Single Cell 3' Reagent Kits v3 (10X Genomics, Pleasanton, California, USA) according to the manufacturer's protocol. The GC suspensions were loaded on the Chromium Single Cell Controller Instrument (10× Genomics, Pleasanton, California, USA) to generate single-cell GEMs (gel beads in emulsions). After that, the RNA from the barcoded cells was subsequently reverse-transcribed to obtain the full-length cDNA. The cDNA libraries were quantiﬁed by Agilent 2100 Bioanalyzer, and subsequently sequenced by OE Biotech Co., Ltd (Shanghai, China) using an BGI-T7 sequencing system (BGI, Shenzhen, China).