title: GSM9119894: SRSF12KO_2nt_brain_rep1; Mus musculus; RNA-Seq
library_construction_protocol: Cells in 6 well plates were washed once with 1x PBS then lysed in 400uL Trizol on plate. To the TRI Reagent lysate, 120 µL of chloroform was added, followed by shaking and centrifugation at 21,000 x g for 15 minutes at 4°C to achieve phase separation. The upper aqueous phase, was carefully transferred to a new tube, mixed with an equal volume of chloroform, and centrifuged again at 21,000 x g for 1 minute at 4°C. RNA was then precipitated by adding 300 mM NaCl and 30 µg of GlycoBlue (AM9516), along with an equal volume of isopropanol, and incubating the mixture overnight at -20°C. The RNA pellet was centrifuged at 21,000 x g for 15 minutes at 4°C, washed with 70% ethanol, and resuspended in RNase-free water. For the last replicate of SRSF12 overexpression analysis, we used Phasemaker tubes to extract RNA. With one exeption (below), all RNA was prepped with the Watchmaker genomics kit with rRNA depletion according to manufacturer's instructions. For the third replicate of SRSF12 overexpression transcriptome analysis, rRNA depletion and library preparation were carried out with the KAPA RNA HyperPrep Kit with RiboErase (HMR) (Roche KK8560) according to manufacturer's instructions.