GSM5668529: Tc0 cells + CoA - mouse 725; Mus musculus; RNA-Seq
RNA was extracted from CD8+ T cells activated for three days in the presence of CoA or Vehicle using Qiagen RNEasy kit. Libraries were prepared using TruSeq Stranded Total RNA kit. Two hundred nanograms from RNA samples were ribosomal RNA depleted using Ribo-zero Gold rRNA beads, following purification the RNA was fragmented. The cleaved RNA fragments were copied into first strand cDNA using reverse transcriptase and random primers. This is followed by second strand cDNA synthesis using RNase H and DNA Polymerase I. A single “A” based were added and adapter ligated followed by purification and enrichment with PCR to create cDNA libraries. Final cDNA libraries were size validated using Agilent Bioanalyzer and concentration validated by qPCR. All libraries were normalized to 10nM and pooled together. 10pM of pooled libraries were loaded onto Illumina cBot for cluster generation. Clustered flow cell was then sequenced Pair-end 100 cycles V3 using Illumina HighSeq 2000 to achieve ~30 million reads per sample.