PolyAdenylated RNA Fragmented (PARFA)-Seq of human cells expressing the vaccinia virus protein F14

Vaccinia virus (VACV) protein F14 is a selective inhibitor of NF-?B. The goal of this experiment was to define the NF-?B-responsive genes that are or are not inhibited by F14 following TNF-a stimulation. T-REx-293 cells inducibly expressing F14 or the empty vector (EV) were treated overnight with doxycycline (to induce virus protein expression). Next day, cells were FBS-starved for 3 h and stimulated with TNF-a for 1 h, 6 h, or left unstimulated. Cells were treated with TNF-a in a staggered manner and all samples were harvested together. Total RNA was prepared by Trizol extraction and processed for PARFA-seq.