Cargando…

Organocatalyst treatment improves variant calling and mutant detection in archival clinical samples

Formalin fixation of biological specimens damages nucleic acids and limits their use in genomic analyses. Previously, we showed that RNA isolation with an organocatalyst (2-amino-5-methylphenyl phosphonic acid, used to speed up reversal of formalin-induced adducts) and extended heated incubation (OR...

Descripción completa

Detalles Bibliográficos
Autores principales: Wehmas, Leah C., Wood, Charles E., Guan, Ping, Gosink, Mark, Hester, Susan D.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group UK 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9021284/
https://www.ncbi.nlm.nih.gov/pubmed/35443772
http://dx.doi.org/10.1038/s41598-022-10301-0
_version_ 1784689777022337024
author Wehmas, Leah C.
Wood, Charles E.
Guan, Ping
Gosink, Mark
Hester, Susan D.
author_facet Wehmas, Leah C.
Wood, Charles E.
Guan, Ping
Gosink, Mark
Hester, Susan D.
author_sort Wehmas, Leah C.
collection PubMed
description Formalin fixation of biological specimens damages nucleic acids and limits their use in genomic analyses. Previously, we showed that RNA isolation with an organocatalyst (2-amino-5-methylphenyl phosphonic acid, used to speed up reversal of formalin-induced adducts) and extended heated incubation (ORGΔ) improved RNA-sequencing data from formalin-fixed paraffin-embedded (FFPE) tissue samples. The primary goal of this study was to evaluate whether ORGΔ treatment improves DNA-sequencing data from clinical FFPE samples. We isolated RNA and DNA ± ORGΔ from paired FFPE and frozen human renal and ovarian carcinoma specimens collected as part of the National Cancer Institute Biospecimen Pre-analytical Variables program. Tumor types were microscopically confirmed from adjacent tissue sections. Following extraction, DNA was fragmented and sequenced and differences were compared between frozen and FFPE sample pairs. Treatment with ORGΔ improved concurrent SNP calls in FFPE DNA compared to non-ORGΔ FFPE samples and enhanced confidence in SNP calls for all FFPE DNA samples, beyond that of matched frozen samples. In general, the concordant SNPs identified in paired frozen and FFPE DNA samples agreed for both genotype and homozygosity vs. heterozygosity of calls regardless of ORGΔ treatment. The increased confidence in ORGΔ FFPE DNA variant calls relative to the matched frozen DNA suggests a novel application of this method. With further optimization, this method may improve quality of DNA-sequencing data in FFPE as well as frozen tissue samples.
format Online
Article
Text
id pubmed-9021284
institution National Center for Biotechnology Information
language English
publishDate 2022
publisher Nature Publishing Group UK
record_format MEDLINE/PubMed
spelling pubmed-90212842022-04-21 Organocatalyst treatment improves variant calling and mutant detection in archival clinical samples Wehmas, Leah C. Wood, Charles E. Guan, Ping Gosink, Mark Hester, Susan D. Sci Rep Article Formalin fixation of biological specimens damages nucleic acids and limits their use in genomic analyses. Previously, we showed that RNA isolation with an organocatalyst (2-amino-5-methylphenyl phosphonic acid, used to speed up reversal of formalin-induced adducts) and extended heated incubation (ORGΔ) improved RNA-sequencing data from formalin-fixed paraffin-embedded (FFPE) tissue samples. The primary goal of this study was to evaluate whether ORGΔ treatment improves DNA-sequencing data from clinical FFPE samples. We isolated RNA and DNA ± ORGΔ from paired FFPE and frozen human renal and ovarian carcinoma specimens collected as part of the National Cancer Institute Biospecimen Pre-analytical Variables program. Tumor types were microscopically confirmed from adjacent tissue sections. Following extraction, DNA was fragmented and sequenced and differences were compared between frozen and FFPE sample pairs. Treatment with ORGΔ improved concurrent SNP calls in FFPE DNA compared to non-ORGΔ FFPE samples and enhanced confidence in SNP calls for all FFPE DNA samples, beyond that of matched frozen samples. In general, the concordant SNPs identified in paired frozen and FFPE DNA samples agreed for both genotype and homozygosity vs. heterozygosity of calls regardless of ORGΔ treatment. The increased confidence in ORGΔ FFPE DNA variant calls relative to the matched frozen DNA suggests a novel application of this method. With further optimization, this method may improve quality of DNA-sequencing data in FFPE as well as frozen tissue samples. Nature Publishing Group UK 2022-04-20 /pmc/articles/PMC9021284/ /pubmed/35443772 http://dx.doi.org/10.1038/s41598-022-10301-0 Text en © This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply 2022 https://creativecommons.org/licenses/by/4.0/Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) .
spellingShingle Article
Wehmas, Leah C.
Wood, Charles E.
Guan, Ping
Gosink, Mark
Hester, Susan D.
Organocatalyst treatment improves variant calling and mutant detection in archival clinical samples
title Organocatalyst treatment improves variant calling and mutant detection in archival clinical samples
title_full Organocatalyst treatment improves variant calling and mutant detection in archival clinical samples
title_fullStr Organocatalyst treatment improves variant calling and mutant detection in archival clinical samples
title_full_unstemmed Organocatalyst treatment improves variant calling and mutant detection in archival clinical samples
title_short Organocatalyst treatment improves variant calling and mutant detection in archival clinical samples
title_sort organocatalyst treatment improves variant calling and mutant detection in archival clinical samples
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9021284/
https://www.ncbi.nlm.nih.gov/pubmed/35443772
http://dx.doi.org/10.1038/s41598-022-10301-0
work_keys_str_mv AT wehmasleahc organocatalysttreatmentimprovesvariantcallingandmutantdetectioninarchivalclinicalsamples
AT woodcharlese organocatalysttreatmentimprovesvariantcallingandmutantdetectioninarchivalclinicalsamples
AT guanping organocatalysttreatmentimprovesvariantcallingandmutantdetectioninarchivalclinicalsamples
AT gosinkmark organocatalysttreatmentimprovesvariantcallingandmutantdetectioninarchivalclinicalsamples
AT hestersusand organocatalysttreatmentimprovesvariantcallingandmutantdetectioninarchivalclinicalsamples