GSM6879746: 293T plus sgRNA, no CMC treatment; Homo sapiens; RNA-Seq
RNA was extracted from harvested cells using the Qiagen RNAeasy kit according to manufacturor's instructions Total RNA was harvested from HEK293T cells  transfected with sgRNA expression plasmid (psU samples), or mock transfected (wt samples),  and polyA-transcripts were selected by magnetic oligodT beads.  Two polyA selections were performed on each sample.  The selected RNA was then either treated with (CMC+) or not (CMC-), adducts other than pseudoU removed by , and the RNAs were reverse transcribed using an RT primer binding to the 3' RNA adaptor. cDNAs were again size selected to enrich for sub-full length RT products. After cleanup, a 3' DNA adaptor was ligated to the cDNA, and the resulting library PCR amplified and sequenced.