Optimisation of laboratory methods for whole transcriptomic RNA analyses in human left ventricular biopsies and blood samples of clinical relevance

Kerrie Ford, Maryam Anwar, Rachael Heys, Eltayeb Mohamed Ahmed, Massimo Caputo, Laurence Game, Barnaby Reeves, Prakash Punjabi, Gianni Angelini, Enrico Petretto, Costanza Emanueli

Research output: Contribution to journalArticle (Academic Journal)

3 Citations (Scopus)
175 Downloads (Pure)

Abstract

This study aimed to optimise techniques for whole transcriptome and small RNA analyses on clinical tissue samples from patients with cardiovascular disease. Clinical samples often represent a particular challenge to extracting RNA of sufficient quality for robust RNA sequencing analysis, and due to availability, it is rarely possible to optimise techniques on the samples themselves. Therefore, we have used equivalent samples from pigs undergoing cardiopulmonary bypass surgery to test different protocols for optimal RNA extraction, and then validated the protocols in human samples. Here we present an assessment of the quality and quantity of RNA obtained using a variety of commercially-available RNA extraction kits on both left ventricular biopsies and blood plasma. RNA extraction from these samples presents different difficulties; left ventricular biopsies are small and fibrous, while blood plasma has a low RNA content. We have validated our optimised extraction techniques on human clinical samples collected as part of the ARCADIA (Association of non-coding RNAs with Coronary Artery Disease and type 2 Diabetes) cohort study, resulting in successful whole transcriptome and small RNA sequencing of human left ventricular tissue.

Original languageEnglish
Article numbere0213685
Number of pages18
JournalPLoS ONE
Volume14
Issue number3
DOIs
Publication statusPublished - 14 Mar 2019

Structured keywords

  • BTC (Bristol Trials Centre)

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  • Projects

    NIHR BRC Cardiovascular

    Angelini, G. D.

    1/04/1731/03/22

    Project: Research, Parent

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