title: GSM9161489: mESC, Auxin 2; Mus musculus; RNA-Seq
library_construction_protocol: Total RNA was purified in triplicates from mESCs with TRIzol (Life Technologies), and RNA integrity (RIN) was corroborated using an Aligent Bioanalyzer. We follow the manufacturer's TruSeq RNA Sample Prep v2 (Illumina, USA) instructions for transcriptome sequencing. The mRNA was purified utilizing poly-T oligo-attached magnetic beads. After purification, the mRNA was fragmented and converted into first-strand cDNA utilizing reverse transcriptase and random primers. The second strand, cDNA, is then synthesized using DNA Polymerase. Actinomycin D was added to prevent DNA-dependent synthesis and improve strand specificity. The cDNA fragments undergo a single 'A' base addition and subsequent adapter ligation. After purification and PCR enrichment, the final cDNA library is suitable for subsequent cluster generation and DNA sequencing. We sequenced the libraries using a NextSeq 500 platform (Illumina) in 75 bp paired end reads format.