DRB-PRO-Seq for measuring Pol II transcriptional elongation rate alterations upon inhibition of analog sensitive CDK12 and/or CDK13

We performed PRO-Seq to determine Pol II elongation rates by measruing the distance of Pol II travels during 25 minutes after Pol II being relased from DRB induced pausing. We allowed Pol II to release in the presence of 1-NM-PP1 or DMSO after DRB being wahsed out in order to assess the effects of CDK12 and/or CDK13 inhibtion on transcriptional elongation rates. Overall design: Cells were treated with 100µM DRB for 4 hours, then DRB was washed out. Washing out of DRB allowed Pol II to elongate, at the meantime, cells were treated with 25 minutes with 1-NM-PP1 or DMSO. Cells were harvested after 25 minutes release, and we measured the distance that Pol II travelled during 25 minutes by PRO-Seq in order to calculate elongation rates.