RNA-Seq of Rattus norvegicus: Adult male PBMNCs
The 5 month old male Sprague Dawley rats, inhoused at the Small animal Facility of Department of Biotechnology, CUSAT were used in this study. Peripheral blood was collected by cardiac puncture and Peripheral Blood Molonuclear cells (PBMNCs) were isolated by density gradient centrifugation using Histopaque 10831. Freshly isolated PBMNCs (Day 0) were resuspended in RNA later solution (Cat No. AM7020, Sigma) and RNA was isolated using RNA isolation kit (Cat.No. 83913, Sigma). The RNA concentration was assessed using Qubit assay. This was followed by RNA enrichment, RNA fragmentation, Random priming, cDNA synthesis and Library Preparation. Library Preparation was performed using NEB Ultra RNA Lib Prep kit.  Library size was assessed using the Agilent 2100 Bioanalyzer, concentration byQubit assay, and final quantitation by qPCR. The sample was then sequenced using Illumina HiSeq platform. The raw data obtained was then taken for RNA Seq analysis.