RNA-seq of Mirounga angustirostris: blubber
Libraries for sequencing were prepared according to TruSeq protocol (TruSeq RNA Sample Prep Kit v2, Illumina, USA). Specifically, mRNA was isolated from total RNA samples using oligo-dT magnetic beads, fragmented and used as template for first-strand cDNA synthesis with random hexamers (SuperScript II Reverse Transcriptase, Invitrogen, USA). After double-stranded (ds) cDNA synthesis and cDNA purification, overhang fragments were end-repaired. This was followed by adenylation of 3_ ends of the blunt-ended fragments and ligation to proprietary adapter oligonucleotides (Illumina, USA). Ligation products were subjected to PCR amplification (10-12 cycles) before library quantification and validation (DNA High Sensitivity Assay, Agilent Technologies, USA).