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Assessing the Performance of Dried-Blood-Spot DNA Extraction Methods in Next Generation Sequencing

An increasing number of newborn screening laboratories in the United States and abroad are moving towards incorporating next-generation sequencing technology, or NGS, into routine screening, particularly for cystic fibrosis. As more programs utilize this technology for both cystic fibrosis and beyon...

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Autores principales: Hendrix, Miyono M., Cuthbert, Carla D., Cordovado, Suzanne K.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7278269/
https://www.ncbi.nlm.nih.gov/pubmed/32514487
http://dx.doi.org/10.3390/ijns6020036
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author Hendrix, Miyono M.
Cuthbert, Carla D.
Cordovado, Suzanne K.
author_facet Hendrix, Miyono M.
Cuthbert, Carla D.
Cordovado, Suzanne K.
author_sort Hendrix, Miyono M.
collection PubMed
description An increasing number of newborn screening laboratories in the United States and abroad are moving towards incorporating next-generation sequencing technology, or NGS, into routine screening, particularly for cystic fibrosis. As more programs utilize this technology for both cystic fibrosis and beyond, it is critical to identify appropriate DNA extraction methods that can be used with dried blood spots that will result in consistent, high-quality sequencing results. To provide comprehensive quality assurance and technical assistance to newborn screening laboratories wishing to incorporate NGS assays, CDC’s Newborn Screening and Molecular Biology Branch designed a study to evaluate the performance of nine commercial or laboratory-developed DNA extraction methods that range from a highly purified column extraction to a crude detergent-based no-wash boil prep. The DNA from these nine methods was used in two NGS library preparations that interrogate the CFTR gene. All DNA extraction methods including the cruder preps performed reasonably well with both library preps. One lower-concentration, older sample was excluded from one of the assay evaluations due to poor performance across all DNA extraction methods. When 84 samples, versus eight, were run on a flow cell, the DNA quality and quantity were more significant variables.
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spelling pubmed-72782692020-06-08 Assessing the Performance of Dried-Blood-Spot DNA Extraction Methods in Next Generation Sequencing Hendrix, Miyono M. Cuthbert, Carla D. Cordovado, Suzanne K. Int J Neonatal Screen Article An increasing number of newborn screening laboratories in the United States and abroad are moving towards incorporating next-generation sequencing technology, or NGS, into routine screening, particularly for cystic fibrosis. As more programs utilize this technology for both cystic fibrosis and beyond, it is critical to identify appropriate DNA extraction methods that can be used with dried blood spots that will result in consistent, high-quality sequencing results. To provide comprehensive quality assurance and technical assistance to newborn screening laboratories wishing to incorporate NGS assays, CDC’s Newborn Screening and Molecular Biology Branch designed a study to evaluate the performance of nine commercial or laboratory-developed DNA extraction methods that range from a highly purified column extraction to a crude detergent-based no-wash boil prep. The DNA from these nine methods was used in two NGS library preparations that interrogate the CFTR gene. All DNA extraction methods including the cruder preps performed reasonably well with both library preps. One lower-concentration, older sample was excluded from one of the assay evaluations due to poor performance across all DNA extraction methods. When 84 samples, versus eight, were run on a flow cell, the DNA quality and quantity were more significant variables. MDPI 2020-04-30 /pmc/articles/PMC7278269/ /pubmed/32514487 http://dx.doi.org/10.3390/ijns6020036 Text en © 2020 by the authors. 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 (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Hendrix, Miyono M.
Cuthbert, Carla D.
Cordovado, Suzanne K.
Assessing the Performance of Dried-Blood-Spot DNA Extraction Methods in Next Generation Sequencing
title Assessing the Performance of Dried-Blood-Spot DNA Extraction Methods in Next Generation Sequencing
title_full Assessing the Performance of Dried-Blood-Spot DNA Extraction Methods in Next Generation Sequencing
title_fullStr Assessing the Performance of Dried-Blood-Spot DNA Extraction Methods in Next Generation Sequencing
title_full_unstemmed Assessing the Performance of Dried-Blood-Spot DNA Extraction Methods in Next Generation Sequencing
title_short Assessing the Performance of Dried-Blood-Spot DNA Extraction Methods in Next Generation Sequencing
title_sort assessing the performance of dried-blood-spot dna extraction methods in next generation sequencing
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7278269/
https://www.ncbi.nlm.nih.gov/pubmed/32514487
http://dx.doi.org/10.3390/ijns6020036
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