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Sample GSM4278307

Query DataSets for GSM4278307

Status

Public on Dec 09, 2020

Title

preOA.H11

Sample type

SRA

Source name

synovium

Organism

Equus caballus

Characteristics

tissue: synovium
status: preOA
disease state: Not affected

Extracted molecule

total RNA

Extraction protocol

Synovial tissue samples were collected arthroscopically from the MCPJ of eleven adult horses before (pre-OA) and 16 weeks after (OA) experimental induction of OA as well as from the sham-operated joints (sham). Synovium samples were placed in RNAlater (Qiagen, Valencia, CA) and stored at -80°C until further processing. Frozen samples were crushed to powder with a mortar and pestle and placed in tubes containing ceramic beads (Precellys® 2ml Hard Tissue Homogenizing Ceramic Beads CK28, Cayman Chemical, Ann Arbor, MI) and TRIzol reagent (Invitrogen, Carlsbad, CA). Mechanical homogenization was performed for cell lysis (Precellys® Minilys, Bertin Corp., Rockville, MD) prior to RNA extraction on spin columns using the RNeasy Micro Kit (Qiagen, Valencia, CA) per manufacturer instructions. Samples were subjected to standard library preparation and sequencing (100 base-pair, paired-end) on an Illumina HiSeq4000 at the University of Illinois Roy J Carver Biotechnology Center.
Libraries were prepared with Illumina's 'TruSeq Stranded mRNAseq Sample Prep kit' and sequenced on an Illumina HiSeq4000. Read 1 aligns to the ANTISENSE strand and Read 2 aligns to the SENSE strand.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 4000

Description

508501RFwk00

Data processing

Quality control on all 28 samples (56 paired-end files) was performed using fastp (version 0.19.5) and summarized using MultiQC (version 1.6)
All fastq files were trimmed using Trimmomatic(version 0.38) in paired end mode to trim any remaining adaptors, then trim leading or trailing bases with quality scores below 28, then discarding a read pair if either read was shorter than 30 bases
Salmon (version 0.11.3) was used to quasi-map to the EquCab3.0 genome assembly (NCBI Annotation Release 103) with --validateMappings --rangeFactorizationBins 4 --gcbias --seqbias options enabled.
R package tximport was used to estimate gene-level counts from transcript-level quantification
Reads were filtered and normalized (TMM) using edgeR, surrogate variables were calculated and counts were corrected using sva, and DE analysis performed using limma-trend
Genome_build: EquCab3.0
Supplementary_files_format_and_content: tab-delimited files with normalized log2(cpm) values for each sample, genes with <1 cpm in 6 samples removed

Submission date

Jan 21, 2020

Last update date

Dec 09, 2020

Contact name

Ann Marie Kemper

E-mail(s)

amkemper@illinois.edu

Organization name

University of Illinois

Street address

1008 W Hazelwood Dr