Yield and Integrity of RNA from Brain Samples are Largely Unaffected by Pre-analytical Procedures
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Yield and Integrity of RNA from Brain Samples are Largely Unaffected by Pre-analytical Procedures Pernille Søs Hovgaard Jensen1 · Maja Johansen1 · Lasse K. Bak2 · Lars Juhl Jensen3 · Christina Kjær1,2 Received: 17 September 2020 / Revised: 13 November 2020 / Accepted: 17 November 2020 © Springer Science+Business Media, LLC, part of Springer Nature 2020
Abstract Gene expression studies are reported to be influenced by pre-analytical factors that can compromise RNA yield and integrity, which in turn may confound the experimental findings. Here we investigate the impact of four pre-analytical factors on brain-derived RNA: time-before-collection, tissue specimen size, tissue collection method, and RNA isolation method. We report no significant differences in RNA yield or integrity between 20 mg and 60 mg tissue samples collected in either liquid nitrogen or the RNAlater stabilizing solution. Isolation of RNA employing the TRIzol reagent resulted in a higher yield compared to isolation via the QIAcube kit while the latter resulted in RNA of slightly better integrity. Keeping brain tissue samples at room temperature for up to 160 min prior to collection and isolation of RNA resulted in no significant difference in yield or integrity. Our findings have significant practical and financial consequences for clinical genomic departments and other laboratory settings performing large-scale routine RNA expression analysis of brain samples. Keywords RNA isolation · RNA Integrity Number (RIN) · Brain tissue · Pre-analytical factors · Gene expression · RNA stability · RNA degradation
Introduction Studies of gene expression require precise and accurate identification and quantification of RNA molecules. Tissue-specific guidelines for a pre-analytical strategy is considered important, as yield and integrity of isolated RNA Pernille Søs Hovgaard Jensen and Maja Johansen have been contributed equally to this publication. Electronic supplementary material The online version of this article (https://doi.org/10.1007/s11064-020-03183-z) contains supplementary material, which is available to authorized users. * Christina Kjær [email protected] 1
Department of Technology, Faculty of Health and Technology, University College Copenhagen, 2200 Copenhagen, Denmark
2
Department of Drug Design and Pharmacology, Faculty of Health and Medical Sciences, University of Copenhagen, 2100 Copenhagen, Denmark
3
Disease Systems Biology Program, Novo Nordisk Foundation Center for Protein Research, Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen, Denmark
are reported to depend on the type of tissue studied [1, 2]. Consequently protection of RNA molecules from enzymatic degradation is important, as it directly impacts RNA levels often leading to significant differences in apparent expression [3, 4]. The RNA Integrity Number (RIN) [5] scale is used to quantify RNA integrity and ranges from 1 to 10, where a higher number indicates more intact RNA. In the following, we use RIN-scores as a
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