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Next-generation sequencing library construction on a surface

BACKGROUND: Next-generation sequencing (NGS) has revolutionized almost all fields of biology, agriculture and medicine, and is widely utilized to analyse genetic variation. Over the past decade, the NGS pipeline has been steadily improved, and the entire process is currently relatively straightforwa...

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Autores principales: Feng, Kuan, Costa, Justin, Edwards, Jeremy S.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5975494/
https://www.ncbi.nlm.nih.gov/pubmed/29848309
http://dx.doi.org/10.1186/s12864-018-4797-4
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author Feng, Kuan
Costa, Justin
Edwards, Jeremy S.
author_facet Feng, Kuan
Costa, Justin
Edwards, Jeremy S.
author_sort Feng, Kuan
collection PubMed
description BACKGROUND: Next-generation sequencing (NGS) has revolutionized almost all fields of biology, agriculture and medicine, and is widely utilized to analyse genetic variation. Over the past decade, the NGS pipeline has been steadily improved, and the entire process is currently relatively straightforward. However, NGS instrumentation still requires upfront library preparation, which can be a laborious process, requiring significant hands-on time. Herein, we present a simple but robust approach to streamline library preparation by utilizing surface bound transposases to construct DNA libraries directly on a flowcell surface. RESULTS: The surface bound transposases directly fragment genomic DNA while simultaneously attaching the library molecules to the flowcell. We sequenced and analysed a Drosophila genome library generated by this surface tagmentation approach, and we showed that our surface bound library quality was comparable to the quality of the library from a commercial kit. In addition to the time and cost savings, our approach does not require PCR amplification of the library, which eliminates potential problems associated with PCR duplicates. CONCLUSIONS: We described the first study to construct libraries directly on a flowcell. We believe our technique could be incorporated into the existing Illumina sequencing pipeline to simplify the workflow, reduce costs, and improve data quality.
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spelling pubmed-59754942018-05-31 Next-generation sequencing library construction on a surface Feng, Kuan Costa, Justin Edwards, Jeremy S. BMC Genomics Methodology Article BACKGROUND: Next-generation sequencing (NGS) has revolutionized almost all fields of biology, agriculture and medicine, and is widely utilized to analyse genetic variation. Over the past decade, the NGS pipeline has been steadily improved, and the entire process is currently relatively straightforward. However, NGS instrumentation still requires upfront library preparation, which can be a laborious process, requiring significant hands-on time. Herein, we present a simple but robust approach to streamline library preparation by utilizing surface bound transposases to construct DNA libraries directly on a flowcell surface. RESULTS: The surface bound transposases directly fragment genomic DNA while simultaneously attaching the library molecules to the flowcell. We sequenced and analysed a Drosophila genome library generated by this surface tagmentation approach, and we showed that our surface bound library quality was comparable to the quality of the library from a commercial kit. In addition to the time and cost savings, our approach does not require PCR amplification of the library, which eliminates potential problems associated with PCR duplicates. CONCLUSIONS: We described the first study to construct libraries directly on a flowcell. We believe our technique could be incorporated into the existing Illumina sequencing pipeline to simplify the workflow, reduce costs, and improve data quality. BioMed Central 2018-05-30 /pmc/articles/PMC5975494/ /pubmed/29848309 http://dx.doi.org/10.1186/s12864-018-4797-4 Text en © The Author(s). 2018 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
spellingShingle Methodology Article
Feng, Kuan
Costa, Justin
Edwards, Jeremy S.
Next-generation sequencing library construction on a surface
title Next-generation sequencing library construction on a surface
title_full Next-generation sequencing library construction on a surface
title_fullStr Next-generation sequencing library construction on a surface
title_full_unstemmed Next-generation sequencing library construction on a surface
title_short Next-generation sequencing library construction on a surface
title_sort next-generation sequencing library construction on a surface
topic Methodology Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5975494/
https://www.ncbi.nlm.nih.gov/pubmed/29848309
http://dx.doi.org/10.1186/s12864-018-4797-4
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