Quality assessment of RNA in long-term storage: The All Our Families biorepository

Nikki L Stephenson; Kylie K Hornaday; Chelsea T A Doktorchik; Andrew W Lyon; Suzanne C Tough; Donna M Slater

doi:10.1371/journal.pone.0242404

Quality assessment of RNA in long-term storage: The All Our Families biorepository

PLoS One. 2020 Dec 1;15(12):e0242404. doi: 10.1371/journal.pone.0242404. eCollection 2020.

Authors

Nikki L Stephenson¹, Kylie K Hornaday², Chelsea T A Doktorchik¹, Andrew W Lyon³, Suzanne C Tough^{1

4}, Donna M Slater^{2

5}

Affiliations

¹ Department of Community Health Sciences, Cumming School of Medicine, University of Calgary, Calgary, Alberta, Canada.
² Department of Physiology and Pharmacology, Cumming School of Medicine, University of Calgary, Calgary, Alberta, Canada.
³ Department of Pathology and Laboratory Medicine, Saskatchewan Health Authority, Saskatoon, Saskatchewan, Canada.
⁴ Department of Paediatrics, Cumming School of Medicine, University of Calgary, Calgary, Alberta, Canada.
⁵ Department of Obstetrics and Gynecology, Cumming School of Medicine, University of Calgary, Calgary, Alberta, Canada.

Abstract

Background: The All Our Families (AOF) cohort study is a longitudinal population-based study which collected biological samples from 1948 pregnant women between May 2008 and December 2010. As the quality of samples can decline over time, the objective of the current study was to assess the association between storage time and RNA (ribonucleic acid) yield and purity, and confirm the quality of these samples after 7-10 years in long-term storage.

Methods: Maternal whole blood samples were previously collected by trained phlebotomists and stored in four separate PAXgene Blood RNA Tubes (PreAnalytiX) between 2008 and 2011. RNA was isolated in 2011 and 2018 using PAXgene Blood RNA Kits (PreAnalytiX) as per the manufacturer's instruction. RNA purity (260/280), as well as RNA yield, were measured using a Nanodrop. The RNA integrity number (RIN) was also assessed from 5-25 and 111-130 months of storage using RNA 6000 Nano Kit and Agilent 2100 BioAnalyzer. Descriptive statistics, paired t-test, and response feature analysis using linear regression were used to assess the association between various predictor variables and quality of the RNA isolated.

Results: Overall, RNA purity and yield of the samples did not decline over time. RNA purity of samples isolated in 2011 (2.08, 95% CI: 2.08-2.09) were statistically lower (p<0.000) than samples isolated in 2018 (2.101, 95% CI: 2.097, 2.104), and there was no statistical difference between the 2011 (13.08 μg /tube, 95% CI: 12.27-13.89) and 2018 (12.64 μg /tube, 95% CI: 11.83-13.46) RNA yield (p = 0.2964). For every month of storage, the change in RNA purity is -0.01(260/280), and the change in RNA yield between 2011 and 2018 is -0.90 μ g / tube. The mean RIN was 8.49 (95% CI:8.44-8.54), and it ranged from 7.2 to 9.5. The rate of change in expected RIN per month of storage is 0.003 (95% CI 0.002-0.004), so while statistically significant, these results are not relevant.

Conclusions: RNA quality does not decrease over time, and the methods used to collect and store samples, within a population-based study are robust to inherent operational factors which may degrade sample quality over time.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Blood Specimen Collection / standards*
Diagnostic Tests, Routine
Female
Humans
Pregnancy
Quality Control
RNA / blood*
RNA / genetics
RNA Stability / genetics*
Specimen Handling / standards*

Substances

RNA

Grants and funding

This study, and the All Our Families Cohort are funded through Alberta Innovates (https://albertainnovates.ca) Interdisciplinary Team Grant (200700595, ST, DS, AL) and the Alberta Children’s Hospital Foundation (http://www.childrenshospital.ab.ca/site/PageNavigator/about/about; RF-ABC001, ST). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.