Evaluation of commercial DNA and RNA extraction methods for high-throughput sequencing of FFPE samples

PloS One
Stine H KresseLeonardo A Meza-Zepeda

Abstract

Nucleic acid material of adequate quality is crucial for successful high-throughput sequencing (HTS) analysis. DNA and RNA isolated from archival FFPE material are frequently degraded and not readily amplifiable due to chemical damage introduced during fixation. To identify optimal nucleic acid extraction kits, DNA and RNA quantity, quality and performance in HTS applications were evaluated. DNA and RNA were isolated from five sarcoma archival FFPE blocks, using eight extraction protocols from seven kits from three different commercial vendors. For DNA extraction, the truXTRAC FFPE DNA kit from Covaris gave higher yields and better amplifiable DNA, but all protocols gave comparable HTS library yields using Agilent SureSelect XT and performed well in downstream variant calling. For RNA extraction, all protocols gave comparable yields and amplifiable RNA. However, for fusion gene detection using the Archer FusionPlex Sarcoma Assay, the truXTRAC FFPE RNA kit from Covaris and Agencourt FormaPure kit from Beckman Coulter showed the highest percentage of unique read-pairs, providing higher complexity of HTS data and more frequent detection of recurrent fusion genes. truXTRAC simultaneous DNA and RNA extraction gave similar outputs as...Continue Reading

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Citations

Feb 29, 2020·Scientific Reports·Margalida Esteva-SociasCristina Villena
Apr 16, 2020·Brazilian Journal of Medical and Biological Research = Revista Brasileira De Pesquisas Médicas E Biológicas·Xinting Lv, Huifei Cheng
Aug 24, 2020·Microbiome·Yensi Flores BuesoMark Tangney
Jul 31, 2020·Diagnostics·Rossella Bruno, Gabriella Fontanini
Mar 1, 2021·BMC Medical Genomics·Carina HeydtSabine Merkelbach-Bruse
Mar 25, 2021·Critical Reviews in Oncology/hematology·Amin TalebiMohammad Amin Kerachian
Apr 23, 2021·Expert Review of Molecular Diagnostics·Svenja Wagener-Ryczek, Roberto Pappesch
May 22, 2021·International Journal of Gynecological Cancer : Official Journal of the International Gynecological Cancer Society·Alba Southern, Mona El-Bahrawy
Jun 21, 2021·Modern Pathology : an Official Journal of the United States and Canadian Academy of Pathology, Inc·Shuko HaradaAlexander C Mackinnon

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Methods Mentioned

BETA
biopsies
nucleic acid extraction
PCR
deamination
Assay
Illumina sequencing

Software Mentioned

Ensembl VEP
Clinvar
ArcherDX
Variant Studio
Archer Analysis
Strelka
COSMIC
Archer Quiver Fusion
Archer
AllPrep

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