organism: Homo sapiens
description: Protocols: All donors provided written informed consent for use of their material, and the collection and use of the samples received Institutional Review Board review and approval. Of the seventeen synovial biopsy samples progressed for RNA-Seq and DNA methylation analyses, fifteen were obtained from knee and two from ankle. Samples were collected from 6 male and 11 female patients and under different medication regimens: no medication, non-steroid anti-inflammatory drugs (NSAID), conventional and biological disease-modifying anti-rheumatic drugs (CDMARD and BDMARD), and combinations thereof. All patients underwent arthroscopic synovial tissue biopsy sampling of an actively inflamed knee joint or ankle. Samples were flash frozen upon collection. Synovial biopsies were disrupted and homogenised in Buffer RLT Plus and β-mercaptoethanol (β-ME) using QIAshredder spin columns followed by simultaneous isolation of DNA and Total RNA using a AllPrep DNA/RNA Mini kit (Qiagen). RNA samples were quantified and their integrity assessed using Qubit RNA Broad Range Assay Kit (Thermo Fisher Scientific) and an Agilent 2100 Bioanalyzer RNA 6000 Nano Kit (Agilent Technologies) respectively. Depending on sample yield, DNA samples were quantified using Qubit DNA BR or Qubit DNA HS kit (Thermo Fisher Scientific). RNA-Seq libraries were prepared using TruSeq® Stranded Total RNA LT - (with Ribo-Zero™ Human/Mouse/Rat) - Set A kit (Illumina), per manufacturer's protocol except for two modifications. First, the time of 'Elution 2 - Frag - Prime' program was reduced from 8 to 6 min to increase the length of the RNA fragments. Secondly, the number of cycles (11 instead of 15) and the time of the initial step in each cycle (30 instead of 10 sec at the first 98°C step) were both modified to minimize PCR-based amplification bias ('PCR' program). Two rounds of qPCR quantification of the final libraries were performed with the KAPA Library Quantification Kit (KAPA Biosystems) on a QuantStudio 12K Flex Real-Time PCR System (Thermo Fisher Scientific).
attributes.tag: ENA-FIRST-PUBLIC
attributes.value: 2019-03-30T17:02:19Z
attributes.tag: ENA-LAST-UPDATE
attributes.value: 2018-04-20T16:41:08Z
attributes.tag: External Id
attributes.value: SAMEA4598851
attributes.tag: INSDC center name
attributes.value: GSK (GlaxoSmithKline)
attributes.tag: INSDC first public
attributes.value: 2019-03-30T17:02:19Z
attributes.tag: INSDC last update
attributes.value: 2018-04-20T16:41:08Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-6684:RA-21
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: common name
attributes.value: human
attributes.tag: disease
attributes.value: rheumatoid arthritis
attributes.tag: individual
attributes.value: P3658488
attributes.tag: institute
attributes.value: T17
attributes.tag: medication
attributes.value: CDMARD
attributes.tag: organism part
attributes.value: synovial membrane
attributes.tag: sample name
attributes.value: E-MTAB-6684:RA-21
attributes.tag: sampling site
attributes.value: knee
attributes.tag: scientific_name
attributes.value: Homo sapiens
attributes.tag: sex
attributes.value: female
attributes.tag: swollen joint count
attributes.value: 0