title: WSX1.D3.1L.p1
organism: Mus musculus
description: Protocols: Studies of antigen-induced arthritis (AIA) were conducted under UK Home Office approvals (PPL-30/1820; PPL-30/2928; PB3E4EE13; PF34A3DC8; PE8BCF782). Mice were challenged (s.c.) with methylated BSA (mBSA; 1 mg/ml emulsified in Complete Freund's Adjuvant) and 160 ng Bordetella pertussis toxin (i.p.). Following an antigen boost with mBSA and CFA (s.c.), inflammatory arthritis was established by intra-articular (i.a.) administration of mBSA (10μl; 10 mg/ml) into the right knee joint.  Synovial tissues were harvested on days 0, 3, and 10 of AIA. Total RNA was isolated from tissue extracts using RNeasy Mini kits (QIAGEN). Libraries were generated from 2-4mg of mRNA following removal of cytoplasmic, mitochondrial, and ribosomal RNA (ribominus transcriptome isolation kit; Ambion). Libraries were prepared using the TrueSeq Stranded Total RNA library Prep kit (Illumina) or RNA-seq kit v2 (Life Technologies; 4475936) and sequenced on an Illumina HiSeq 4000 platform. RNA was extracted utilising QIAshredders (Qiagen) and RNeasy Mini Kits (Qiagen). On-column DNase digestion (Qiagen) was utilised to remove contaminating genomic DNA. Single-stranded cDNA was synthesized, processed and labelled sequentially using the Ambion® WT (Whole Transcript) Expression Kit, the Affymetrix® Genechip® Poly-A RNA Control Kit and the Terminal Labelling kit.