Comparison of whole transcriptome sequencing of fresh, frozen, and formalin-fixed, paraffin-embedded cardiac tissue

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The use of fresh tissue for molecular studies is preferred but often impossible. Instead, frozen or formalin-fixed, paraffin-embedded (FFPE) tissues are widely used and constitute valuable resources for retrospective studies. We assessed the utility of cardiac tissue stored in different ways for gene expression analyses by whole transcriptome sequencing of paired fresh, frozen, and FFPE tissues. RNA extracted from FFPE was highly degraded. Sequencing of RNA from FFPE tissues yielded higher proportions of intronic and intergenic reads compared to RNA from fresh and frozen tissues. The global gene expression profiles varied with the storage conditions, particularly mitochondrial and long non-coding RNAs. However, we observed high correlations among protein-coding transcripts (ρ > 0.94) with the various storage conditions. We did not observe any significant storage effect on the allele-specific gene expression. However, FFPE had statistically significantly (p < 0.05) more discordant variant calls compared to fresh and frozen tissue. In conclusion, we found that frozen and FFPE tissues can be used for reliable gene expression analyses, provided that proper quality control is performed and caution regarding the technical variability is withheld.

Original languageEnglish
Article numbere0283159
JournalPLoS ONE
Issue number3
Number of pages15
Publication statusPublished - 2023

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Copyright: © 2023 Jacobsen et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

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