RNA-seq of mouse fibroblast-like synoviocytes (FLS) isolated from mice with collagen-induced arthritis (CIA) and naive littermates
RNA-seq of mouse fibroblast-like synoviocytes (FLS) isolated from mice with collagen-induced arthritis (CIA) and naive littermates
Paws were dissected to remove skin and separate joints, then incubated with collagenase IV (10 mg/ml, Sigma C5138) in DMEM containing 10% FBS and 1% penicillin-streptomycin at 37ºC with shaking for 1-1.5 hours.  FLS cells were selected using the EasySep Release Mouse PE Positive Selection kit (Stemcell, 17656) and a Podoplanin-PE conjugated antibody (BioLegend 127407). FLS cell pellets were lysed in buffer RLT Plus supplemented with DTT, and RNA isolated using the RNeasy Micro Plus kit (Qiagen) according to the manufacturer's instructions.   Sequencing libraries were generated using the Stranded mRNA Prep Ligation kit (Illumina, Inc) according to the manufacturer's protocol.  Library fragment size and quality was assayed using a 4200 TapeStation (Agilent Technologies).