HDFn-KLF4-d3-RNAseq-2
HDFn cells were grown in Medium 106 (Thermo Fisher Scientific, M-106-500) to 33-66% confluency in 6-well plates and spinfected (1 hour at 2000 rpm) with KLF4 lentivirus (3X copy number). The spinfection procedure was repeated 24 hours later and fresh media was added at 48 hours. Cells were grown for 3 days after the initial infection, collected by trypsinization, and frozen at -20C. Total RNA was isolated for RNA-seq analysis from the cell pellet using the RNAqueous-Micro Total RNA Isolation Kit (Thermo Fisher Scientific, AM1931). Samples were treated with DNase I for 20 minutes at room temperature. RNA-seq library preparation and sequencing was performed at the Vanderbilt VANTAGE core. Stranded RNA-seq libraries were prepared from total RNA using poly-A enrichment and the TruSeq RNA Library Preparation Kit (Illumina). Libraries were sequenced on an Illumina NovaSeq 6000 or NextSeq 500 using a paired-end 150 bp protocol.