Cord blood CD4 T cells response to PMA/Ionomycin - bulk RNA-Seq
RNA from CD4 T cell pellets (n=3 lean, 3 obese) following PMA stimulation was extracted using mRNeasy kit (Qiagen, Valencia CA). RNA concentration and integrity were determined using Agilent 2100 Bioanalyzer. Libraries were generated using TruSeq Stranded Total RNA-Seq kit (Illumina, San Diego CA). Following rRNA depletion, mRNA was fragmented, converted to double stranded cDNA and adapter ligated. Fragments were enriched by PCR amplification and purified. Library concentrations were measured using Qubit; library quality and peak sizes were assessed on the Bioanalyzer. Libraries were multiplexed and sequenced on the HiSeq4000 platform to yield an average 20 million 100 bp single end reads.