mRNA-seq whole transcriptome profiling of fresh frozen versus archived fixed tissues

BMC Genomics
Noa Bossel Ben-MosheMaya Dadiani

Abstract

The main bottleneck for genomic studies of tumors is the limited availability of fresh frozen (FF) samples collected from patients, coupled with comprehensive long-term clinical follow-up. This shortage could be alleviated by using existing large archives of routinely obtained and stored Formalin-Fixed Paraffin-Embedded (FFPE) tissues. However, since these samples are partially degraded, their RNA sequencing is technically challenging. In an effort to establish a reliable and practical procedure, we compared three protocols for RNA sequencing using pairs of FF and FFPE samples, both taken from the same breast tumor. In contrast to previous studies, we compared the expression profiles obtained from the two matched sample types, using the same protocol for both. Three protocols were tested on low initial amounts of RNA, as little as 100 ng, to represent the possibly limited availability of clinical samples. For two of the three protocols tested, poly(A) selection (mRNA-seq) and ribosomal-depletion, the total gene expression profiles of matched FF and FFPE pairs were highly correlated. For both protocols, differential gene expression between two FFPE samples was in agreement with their matched FF samples. Notably, although express...Continue Reading

References

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Citations

Oct 23, 2019·Nature Methods·Shalom Hillel RothEli Eisenberg
Dec 26, 2019·PeerJ·Macarena ArroyoM Gonzalo Claros
Dec 20, 2020·Clinical Genetics·Agata SzymiczekMohammad R Akbari
Apr 19, 2021·European Journal of Cancer : Official Journal for European Organization for Research and Treatment of Cancer (EORTC) [and] European Association for Cancer Research (EACR)·Maria KuksinLoïc Verlingue
Dec 12, 2020·Arthritis & Rheumatology·Jessica L TurnierJ Michelle Kahlenberg

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Methods Mentioned

BETA
RNA-seq
biopsies
nucleic acid isolation
PCR

Software Mentioned

ngs
RiboZero
mRNA
seq
HTSeq
TopHat

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