title: BMDM-Control-4h
design: Total RNA was extracted using TRIzol® (Life Technologies, Carlsbad, CA, USA) according to the manufacturer’s protocol. For RNA-Seq, RNA libraries were created from each group using the NEBNext® Ultra™ Directional RNA Library preparation kit from Illumina® Illumina, San Diego, CA, USA). The first step in the workflow involved the removal of ribosomal RNA using the RNAMius™ Transcriptome Isolation kit (Life Technologies, Carlsbad, CA, USA).  Following purification, total RNA was fragmented into small pieces using divalent cations at elevated temperature. The cleaved RNA fragments were copied into first-strand cDNA using reverse transcriptase and random primers, followed by second-strand cDNA synthesis using DNA polymerase I and RNase H. The cDNA fragments were then processed through an end-repair reaction by the addition of a single ‘A’ base, followed by ligation of the adapters. The products of these reactions were then purified and enriched by PCR to create the final cDNA library. The cDNA fragments were sequenced using the Illumina HiSeq2500 (101 cycles PE lane) (NICEM).