Time-series RNA-Seq upon actinomycin D (ActD) treatment

PrEC and C4-2 cells were treated with transcription inhibitor actinomycin D (ActD, Sigma) at a concentration of 5 μg/ml for 0, 15 min, 1 h and 2 h. RNA samples were then isolated using RNeasy Plus Mini Kit (Qiagen) and sent to BGI for library preparation and follow-up sequencing. Human primary total T cells were isolated from the blood sample of three donors from Gulf Coast Regional Blood Center using Ficoll density gradient method. The cells were collected and washed with PBS + 0.5% FBS and taken into culture. After isolation, the cells were activated with anti-CD3/CD28 microbeads (Thermo Fisher) for 24 h. For transcription inhibition, the cultured cells were treated with ActD (10 μg/ml), for four time points, 0 h (immediately after ActD treatment), 15 min, 1 h and 2 h. After treatment, RNA was isolated by RNeasy kit (Qiagen). The total RNA-seq library construction was performed by RNA core.