RNA-Seq of CFT073 cultured in RPMI 1640 medium
The mRNA was fragmented using divalent cations under elevated temperatures and synthesize double-stranded cDNA. The ends of these cDNA fragments were end repaired and A-tailed. The A-tailed fragments were then purified and ligated to Illumina adapters. These products were subsequently loaded into 2% low melting point agarose gels. The cDNA fragments between 200 and 250bp in size were selected and purified. After purification, a PCR assay using the obtained cDNA was performed, and the PCR products were purified. Cluster formation, primer hybridization and single-end 36-cycle sequencing was performed.