LPS (16h) Stimulated Splenic Macrophages from CHD animals
Total RNA was isolated from CD14+ cells from the spleen using mRNeasy kit (Qiagen, Valencia CA) and quality assessed using Agilent 2100 Bioanalyzer. Following ribosomal RNA depletion using Ribo-Gone rRNA removal kit (Clontech, Mountain View CA), libraries were generated using SMARTer Stranded RNA-Seq kit. Briefly, rRNA depleted RNA was fragmented, converted to double stranded cDNA and ligated to adapters. The roughly 300bp-long fragments were then amplified by PCR and selected by size exclusion. Libraries were multiplexed using unique barcodes to facilitate sequencing of several biological samples. Following quality control for size, quality, and concentrations, libraries of 100bp read lengths were sequenced on single-end mode.