GSM7016827: 8-1 KO veh_3394; Mus musculus; RNA-Seq
RNA was extracted from heart tissue, CMs or CFs using either RNeasy kit (Qiagen) The total RNA concentration was determined with the NanopDrop 1000 spectrophotometer (NanoDrop, Wilmington, DE) and RNA quality assessed with the Agilent Bioanalyzer (Agilent, Santa Clara, CA). The TruSeq Stranded mRNA Sample Preparation Kit (Illumina, San Diego, CA) was used for next generation sequencing library construction per manufacturer's protocols. Briefly, mRNA was purified from 200ng total RNA with oligo-dT magnetic beads and fragmented. First-strand cDNA synthesis was performed with random hexamer priming followed by second-strand cDNA synthesis using dUTP incorporation for strand marking. End repair and 3` adenylation was then performed on the double stranded cDNA. Illumina adaptors were ligated to both ends of the cDNA and amplified with PCR primers specific to the adaptor sequences to generate cDNA amplicons of approximately 200-500bp in size. Amplified libraries were sequenced using the NextSeq 550 sequencer (Illumina, San Diego, CA) on the High flow cell to obtain approximately 25 to 30 million reads per sample.