GSM8448926: Old_GF; Drosophila melanogaster; RNA-Seq
Fly brain dissection and tissue dissociation were conducted before 11 AM. Fly brains were dissected in ice-cold Schneider's Drosophila medium (Gibco), with twenty brains pooled for each sample. The dissected brains were immediately incubated with 1.2 mg/mL Dispase (Sigma-Aldrich) and 0.6 mg/mL Collagenase (Invitrogen) in a shaking incubator (Eppendorf) at 25°C and 500 rpm for 75 minutes. Following the incubation, the samples were washed three times with ice-cold Dulbecco's phosphate-buffered saline (DPBS, Gibco) and resuspended in 50 µm DPBS with 0.04% bovine serum albumin (BSA, Sigma). The cell suspension was then passed through a 35 µm cell strainer (Falcon) to remove debris. Cell number and viability were assessed simultaneously using an epifluorescence microscope (Leica) and an Automated Cell Counter (LUNA) to ensure the cell viability to exceed 90%. Single-cell libraries were prepared using the Chromium Single Cell 3' Reagent Kit v2 (10x Genomics) in accordance with the manufacturer's manual.