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Combing of Genomic DNA from Droplets Containing Picograms of Material
[Image: see text] Deposition of linear DNA molecules is a critical step in many single-molecule genomic approaches including DNA mapping, fiber-FISH, and several emerging sequencing technologies. In the ideal situation, the DNA that is deposited for these experiments is absolutely linear and uniform...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Chemical Society
2015
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4344373/ https://www.ncbi.nlm.nih.gov/pubmed/25561163 http://dx.doi.org/10.1021/nn5063497 |
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author | Deen, Jochem Sempels, Wouter De Dier, Raf Vermant, Jan Dedecker, Peter Hofkens, Johan Neely, Robert K. |
author_facet | Deen, Jochem Sempels, Wouter De Dier, Raf Vermant, Jan Dedecker, Peter Hofkens, Johan Neely, Robert K. |
author_sort | Deen, Jochem |
collection | PubMed |
description | [Image: see text] Deposition of linear DNA molecules is a critical step in many single-molecule genomic approaches including DNA mapping, fiber-FISH, and several emerging sequencing technologies. In the ideal situation, the DNA that is deposited for these experiments is absolutely linear and uniformly stretched, thereby enabling accurate distance measurements. However, this is rarely the case, and furthermore, current approaches for the capture and linearization of DNA on a surface tend to require complex surface preparation and large amounts of starting material to achieve genomic-scale mapping. This makes them technically demanding and prevents their application in emerging fields of genomics, such as single-cell based analyses. Here we describe a simple and extremely efficient approach to the deposition and linearization of genomic DNA molecules. We employ droplets containing as little as tens of picograms of material and simply drag them, using a pipet tip, over a polymer-coated coverslip. In this report we highlight one particular polymer, Zeonex, which is remarkably efficient at capturing DNA. We characterize the method of DNA capture on the Zeonex surface and find that the use of droplets greatly facilitates the efficient deposition of DNA. This is the result of a circulating flow in the droplet that maintains a high DNA concentration at the interface of the surface/solution. Overall, our approach provides an accessible route to the study of genomic structural variation from samples containing no more than a handful of cells. |
format | Online Article Text |
id | pubmed-4344373 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2015 |
publisher | American Chemical Society |
record_format | MEDLINE/PubMed |
spelling | pubmed-43443732015-03-02 Combing of Genomic DNA from Droplets Containing Picograms of Material Deen, Jochem Sempels, Wouter De Dier, Raf Vermant, Jan Dedecker, Peter Hofkens, Johan Neely, Robert K. ACS Nano [Image: see text] Deposition of linear DNA molecules is a critical step in many single-molecule genomic approaches including DNA mapping, fiber-FISH, and several emerging sequencing technologies. In the ideal situation, the DNA that is deposited for these experiments is absolutely linear and uniformly stretched, thereby enabling accurate distance measurements. However, this is rarely the case, and furthermore, current approaches for the capture and linearization of DNA on a surface tend to require complex surface preparation and large amounts of starting material to achieve genomic-scale mapping. This makes them technically demanding and prevents their application in emerging fields of genomics, such as single-cell based analyses. Here we describe a simple and extremely efficient approach to the deposition and linearization of genomic DNA molecules. We employ droplets containing as little as tens of picograms of material and simply drag them, using a pipet tip, over a polymer-coated coverslip. In this report we highlight one particular polymer, Zeonex, which is remarkably efficient at capturing DNA. We characterize the method of DNA capture on the Zeonex surface and find that the use of droplets greatly facilitates the efficient deposition of DNA. This is the result of a circulating flow in the droplet that maintains a high DNA concentration at the interface of the surface/solution. Overall, our approach provides an accessible route to the study of genomic structural variation from samples containing no more than a handful of cells. American Chemical Society 2015-01-05 2015-01-27 /pmc/articles/PMC4344373/ /pubmed/25561163 http://dx.doi.org/10.1021/nn5063497 Text en Copyright © 2015 American Chemical Society This is an open access article published under an ACS AuthorChoice License (http://pubs.acs.org/page/policy/authorchoice_termsofuse.html) , which permits copying and redistribution of the article or any adaptations for non-commercial purposes. |
spellingShingle | Deen, Jochem Sempels, Wouter De Dier, Raf Vermant, Jan Dedecker, Peter Hofkens, Johan Neely, Robert K. Combing of Genomic DNA from Droplets Containing Picograms of Material |
title | Combing of Genomic DNA from Droplets Containing Picograms of Material |
title_full | Combing of Genomic DNA from Droplets Containing Picograms of Material |
title_fullStr | Combing of Genomic DNA from Droplets Containing Picograms of Material |
title_full_unstemmed | Combing of Genomic DNA from Droplets Containing Picograms of Material |
title_short | Combing of Genomic DNA from Droplets Containing Picograms of Material |
title_sort | combing of genomic dna from droplets containing picograms of material |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4344373/ https://www.ncbi.nlm.nih.gov/pubmed/25561163 http://dx.doi.org/10.1021/nn5063497 |
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