title: GSM6051416: 19340X2 Female, 6-week Adrenal Gland, SF1-Cre+;Znrf3Fx/Fx; Mus musculus; RNA-Seq
library_construction_protocol: Tissues were finely chopped and digested enzymatically using enzymes and reagents from the Neural Tissue Dissociation Kit (Mitenyi Biotec, 130-092-628). All steps were carried out at 4oC, including centrifugation. All tubes and pipette tips used to handle cell suspensions were pre-coated with 3% BSA in HBSS to prevent cell loss. During the dissociation, the cell suspension was gently agitated with mechanical pipetting every 10 minutes and visually assessed under a stereo microscope until the tissue was fully digested. The suspension was then filtered through 70-micron filters to obtain a single cell suspension and enzymes were neutralized using HBSS containing 10% Fetal Bovine Serum (FBS). Red blood cells were removed using Red Blood Cell Lysis buffer (Roche, 11814389001) according to manufacturer guidelines, and the cells were washed twice in HBSS containing 2% FBS before counting using a Countess Automated Cell Counter (Thermo Fisher). Immediately following dissociation, cells were stained with 0.1 mg/mL DAPI and 5mM Vybrant DyeCycle Ruby (Invitrogen, V10309). Viable Ruby-positive, DAPI-negative single cells were sorted based on fluorescent properties using the BD FACSAria III Cell Sorter at the University of Utah Flow Cytometry Shared Resource. Cells were collected in 1XPBS (calcium- and magnesium-free) containing 0.04% BSA and kept on ice. Chromium Next GEM Single Cell 3' Gene Expression Library Construction Kit v3.1