title: Combing of genomic DNA from droplets containing picograms of material.
abstract: 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.
authors.affiliation: Department of Chemistry, KU Leuven , Celestijnenlaan 200F, Heverlee 3001, Belgium.
authors.forename: Jochem
authors.initials: J
authors.lastname: Deen
authors.forename: Wouter
authors.initials: W
authors.lastname: Sempels
authors.forename: Raf
authors.initials: R
authors.lastname: De Dier
authors.forename: Jan
authors.initials: J
authors.lastname: Vermant
authors.forename: Peter
authors.initials: P
authors.lastname: Dedecker
authors.forename: Johan
authors.initials: J
authors.lastname: Hofkens
authors.forename: Robert K
authors.initials: RK
authors.lastname: Neely
mesh_terms: D015894:Genome, Human; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D061925:Immobilized Nucleic Acids / Q000737:chemistry*; D055595:Mechanical Phenomena; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D011108:Polymers / Q000737:chemistry; D013499:Surface Properties
chemical_list: D061925:Immobilized Nucleic Acids; D011108:Polymers
keywords: DNA deposition; DNA mapping; coffee ring effect; fiber-FISH; molecular combing; rolling droplet; single-molecule imaging