GSM8330287
RNA was isolated from 200 µl of cells using the RNAsnap procedure (Stead et al. 2012). Two volumes of RNAprotect (Qiagen) were added and cells were incubated for 5 min. Cells were pelleted at full-speed for 20 min at 4 °C and the supernatant was removed. The pellet was resuspended in 100 µl SNAP buffer (0.025% SDS, 18 mM EDTA, 1% β-mercaptoethanol, 95% formamide (RNA-grade)). Samples were incubated for 7 min at 95 °C, cell debris was pelleted at full-speed for 5 min at room-temperature, and the supernatant was transferred to a new tube. 1.5 volumes of ethanol was added to the supernatant and the sample was mixed by pipetting. The sample was loaded onto a miRNeasy mini column (Qiagen) two times and spun at full-speed for 20 s at room-temperature. Columns were washed two times with 500 µ