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Investigation of Different Library Preparation and Tissue of Origin Deconvolution Methods for Urine and Plasma cfDNA Methylome Analysis
Methylation sequencing is a promising approach to infer the tissue of origin of cell-free DNA (cfDNA). In this study, a single- and a double-stranded library preparation approach were evaluated with respect to their technical biases when applied on cfDNA from plasma and urine. Additionally, tissue o...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10417284/ https://www.ncbi.nlm.nih.gov/pubmed/37568867 http://dx.doi.org/10.3390/diagnostics13152505 |
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author | Kueng, Nicholas Sidler, Daniel Banz, Vanessa Largiadèr, Carlo R. Ng, Charlotte K. Y. Amstutz, Ursula |
author_facet | Kueng, Nicholas Sidler, Daniel Banz, Vanessa Largiadèr, Carlo R. Ng, Charlotte K. Y. Amstutz, Ursula |
author_sort | Kueng, Nicholas |
collection | PubMed |
description | Methylation sequencing is a promising approach to infer the tissue of origin of cell-free DNA (cfDNA). In this study, a single- and a double-stranded library preparation approach were evaluated with respect to their technical biases when applied on cfDNA from plasma and urine. Additionally, tissue of origin (TOO) proportions were evaluated using two deconvolution methods. Sequencing cfDNA from urine using the double-stranded method resulted in a substantial within-read methylation bias and a lower global methylation (56.0% vs. 75.8%, p ≤ 0.0001) compared to plasma cfDNA, both of which were not observed with the single-stranded approach. Individual CpG site-based TOO deconvolution resulted in a significantly increased proportion of undetermined TOO with the double-stranded method (urine: 32.3% vs. 1.9%; plasma: 5.9% vs. 0.04%; p ≤ 0.0001), but no major differences in proportions of individual cell types. In contrast, fragment-level deconvolution led to multiple cell types, with significantly different TOO proportions between the two methods. This study thus outlines potential limitations of double-stranded library preparation for methylation analysis of cfDNA especially for urinary cfDNA. While the double-stranded method allows jagged end analysis in addition to TOO analysis, it leads to significant methylation bias in urinary cfDNA, which single-stranded methods can overcome. |
format | Online Article Text |
id | pubmed-10417284 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-104172842023-08-12 Investigation of Different Library Preparation and Tissue of Origin Deconvolution Methods for Urine and Plasma cfDNA Methylome Analysis Kueng, Nicholas Sidler, Daniel Banz, Vanessa Largiadèr, Carlo R. Ng, Charlotte K. Y. Amstutz, Ursula Diagnostics (Basel) Brief Report Methylation sequencing is a promising approach to infer the tissue of origin of cell-free DNA (cfDNA). In this study, a single- and a double-stranded library preparation approach were evaluated with respect to their technical biases when applied on cfDNA from plasma and urine. Additionally, tissue of origin (TOO) proportions were evaluated using two deconvolution methods. Sequencing cfDNA from urine using the double-stranded method resulted in a substantial within-read methylation bias and a lower global methylation (56.0% vs. 75.8%, p ≤ 0.0001) compared to plasma cfDNA, both of which were not observed with the single-stranded approach. Individual CpG site-based TOO deconvolution resulted in a significantly increased proportion of undetermined TOO with the double-stranded method (urine: 32.3% vs. 1.9%; plasma: 5.9% vs. 0.04%; p ≤ 0.0001), but no major differences in proportions of individual cell types. In contrast, fragment-level deconvolution led to multiple cell types, with significantly different TOO proportions between the two methods. This study thus outlines potential limitations of double-stranded library preparation for methylation analysis of cfDNA especially for urinary cfDNA. While the double-stranded method allows jagged end analysis in addition to TOO analysis, it leads to significant methylation bias in urinary cfDNA, which single-stranded methods can overcome. MDPI 2023-07-27 /pmc/articles/PMC10417284/ /pubmed/37568867 http://dx.doi.org/10.3390/diagnostics13152505 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Brief Report Kueng, Nicholas Sidler, Daniel Banz, Vanessa Largiadèr, Carlo R. Ng, Charlotte K. Y. Amstutz, Ursula Investigation of Different Library Preparation and Tissue of Origin Deconvolution Methods for Urine and Plasma cfDNA Methylome Analysis |
title | Investigation of Different Library Preparation and Tissue of Origin Deconvolution Methods for Urine and Plasma cfDNA Methylome Analysis |
title_full | Investigation of Different Library Preparation and Tissue of Origin Deconvolution Methods for Urine and Plasma cfDNA Methylome Analysis |
title_fullStr | Investigation of Different Library Preparation and Tissue of Origin Deconvolution Methods for Urine and Plasma cfDNA Methylome Analysis |
title_full_unstemmed | Investigation of Different Library Preparation and Tissue of Origin Deconvolution Methods for Urine and Plasma cfDNA Methylome Analysis |
title_short | Investigation of Different Library Preparation and Tissue of Origin Deconvolution Methods for Urine and Plasma cfDNA Methylome Analysis |
title_sort | investigation of different library preparation and tissue of origin deconvolution methods for urine and plasma cfdna methylome analysis |
topic | Brief Report |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10417284/ https://www.ncbi.nlm.nih.gov/pubmed/37568867 http://dx.doi.org/10.3390/diagnostics13152505 |
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