Improving Transcriptome Fidelity Following Synovial Tissue Disaggregation.

ABSTRACT:

Objective

To improve the fidelity of the cellular transcriptome of disaggregated synovial tissue for applications such as single-cell RNA sequencing (scRNAseq) by modifying the disaggregation technique.

Methods

Osteoarthritis (OA) and rheumatoid arthritis (RA) synovia were collected at arthroplasty. RNA was extracted from intact or disaggregated replicate pools of tissue fragments. Disaggregation was performed with either a proprietary protease, Liberase TL (Lib) as a reference method, Liberase TL with an RNA polymerase inhibitor flavopyridol (Flavo), or a cold digestion with subtilisin A (SubA). qPCR on selected markers and RNAseq were used to compare disaggregation methods using the original intact tissue as reference.

Results

Disaggregated cell yield and viability

SUBMITTER: Boyle DL 

PROVIDER: S-EPMC9400013 | biostudies-literature | 2022

REPOSITORIES: biostudies-literature