GSM9137018: no heat shock, -Dox ctl, 4sU RNA-seq, R1; Homo sapiens; RNA-Seq
Cells were treated with 1mM 4sU for 20 min prior to harvesting, or the last 20 min of heat shock. Total RNA was extracted and 4sU labeled RNA was conjugated to MSEA-XX-Biotin and isolated using the Miltenyi uMACS streptavidin kit. Libraries for Illumina sequencing were prepared from 4sU RNA by the University of Colorado Anschutz Genomics and Microarray Core facility first depleting rRNA with the Lexogen RiboCop rRNA depletion kit and then libraries were prepared using the TECAN Universal Plus mRNA-seq library prep kit, skipping the poly-A selection (HS 4sU RNA samples) or Zymo-seq RiboFree Total RNA (N-RBBP6 4sU RNA samples). Sequencing was performed using an Illumina NovaSEQ 6000 or NovaSEQ X, obtaining 2 × 150 bp reads.