RNA sequencing of PIK3CA-WT/WT and PIK3CA-WT/H1047R primary mouse embryonic fibroblasts

Gene targeting was carried out by Taconic Artemis (Cologne, Germany) to introduce a point mutation resulting in conversion of CAT (H) to CGT (R) in the C-terminal p110a kinase domain, and a neo selection cassette flanked by frt sites (publication doi: 10.1038/s41467-017-02002-4). MEFs were made from PIK3CA-H1047R/WT and PIK3CA-WT/WT E13.5 embryos (4 independent MEFs per genotype), treated with 1?µM 4-OHT for one day and gathered 48h after the start of 4-OHT induction were subjected to mRNA sequencing to determine changes in gene expression upon short term PIK3CA activation in the absence of other transforming mutations. PIK3CA-H1047R/WT MEFS exhibited altered expression of 71 genes (q cutoff <0.05). Overall design: 4 independent MEFs (WT/WT, WT/H1047R). All cultures were collected 48 hours after 4-OHT induction of flp recombinase mediated PIK3CA mutation. Cells were grown in full media (doi: 10.1038/srep39985). The library was prepared by The Barts and the London School of Medicine and Dentistry Genome Centre at Barts Cancer Centre, using an Illumina TruSeq Standard mRNA Library Prep Kit and sequenced on a Illumina Next Generation Sequencer.