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Handling of long DNA - applications and polymer physics

Kühnlein, Alexandra LU (2016) FYSM60 20152
Solid State Physics
Department of Physics
Abstract
This thesis provides a proof of principle that deterministic lateral displacement (DLD) can be used to spatially separate DNA molecules by size. In order to achieve high separation quality, the pressure range, i.e. flow velocity has to be chosen carefully. The experiments were conducted in DLD devices fabricated in PDMS using soft lithography methods. Fluorescently labeled DNA molecules were transported through the device by pressure driven flow. The separation results were determined from and the separation result was recorded optically at the outlet. Both tested devices, with a critical diameter Dc of 0.64 μm and 0.75 μm respectively, showed capable of separating <10 kbp from 48.5 kbp DNA molecules with a separation quality of ~94%.
Popular Abstract
DNA is the blueprint for life and contains all necessary information to ’build’ an organism. Therefore, DNA might be considered as the most important biologically relevant molecule. However, even though DNA is such a substantial molecule, the research methods and techniques regarding DNA are often surprisingly limited which is due to the complicated handling of the long molecule. This is also the case for DNA sorting methods, i.e the
process of separating different DNA molecules from each other dependent on characteristic factors like weight or length. Sorting is especially important as pre-processing method for other established analysis techniques. Currently, the length-based sorting of DNA molecules is predominantly done using... (More)
DNA is the blueprint for life and contains all necessary information to ’build’ an organism. Therefore, DNA might be considered as the most important biologically relevant molecule. However, even though DNA is such a substantial molecule, the research methods and techniques regarding DNA are often surprisingly limited which is due to the complicated handling of the long molecule. This is also the case for DNA sorting methods, i.e the
process of separating different DNA molecules from each other dependent on characteristic factors like weight or length. Sorting is especially important as pre-processing method for other established analysis techniques. Currently, the length-based sorting of DNA molecules is predominantly done using electrophoresis, for which a huge DNA sample volume with roughly a million molecules is required.
Microfluidics is a relatively new, expanding field in biophysics which deals with fluids on the microliter and micrometer scale. Microfluidics are contrasted by macrofluidics which is our ’normal’ perception of fluids. An easy example which points out the difference is the surface tension of fluids. In every day life we experience it only as a side effect, but when dealing with cellular volumes, typically 1 μl which is one millionth of a liter, the surface tension wins over gravity.
One application of microfluidics is the deterministic lateral displacement method, also called DLD, which was introduced by Richard Huang et al. in 2004. It is based on a micrometer-sized channel in which small obstacles are placed. If a fluid which also contains small particles flows through this obstacle-larded channel, the track of the particles vary dependent on their size: while very small particles follow the fluid flow and pass straight through the device, slightly larger particles get deflected to one of the channel walls. Thus, differently sized particles can be distinguished at the end of the channel as they get spatially separated.
During the following master thesis, the sorting of DNA molecules dependent on their length was investigated. For this, the described DLD method was applied to DNA molecules in solution. It was found that DLD - if used under correct conditions - is an appropriate method to sort DNA molecules dependent on their length.
This finding can be used in sample preparation and processing steps for DNA related experiments. Furthermore, since this is a microfluidic approach it allows DNA length separation for extremely small sample sizes, with only a couple of hundreds of molecules. Especially considering advancing single molecule analysis techniques and personalized medicine approaches, a cut-down sample volume will be crucial in future developments. (Less)
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author
Kühnlein, Alexandra LU
supervisor
organization
course
FYSM60 20152
year
type
H2 - Master's Degree (Two Years)
subject
keywords
Biophysics, microfluidics, DLD, DNA, sorting
language
English
id
8888414
date added to LUP
2016-08-16 09:49:39
date last changed
2016-08-16 09:49:39
@misc{8888414,
  abstract     = {This thesis provides a proof of principle that deterministic lateral displacement (DLD) can be used to spatially separate DNA molecules by size. In order to achieve high separation quality, the pressure range, i.e. flow velocity has to be chosen carefully. The experiments were conducted in DLD devices fabricated in PDMS using soft lithography methods. Fluorescently labeled DNA molecules were transported through the device by pressure driven flow. The separation results were determined from and the separation result was recorded optically at the outlet. Both tested devices, with a critical diameter Dc of 0.64 μm and 0.75 μm respectively, showed capable of separating <10 kbp from 48.5 kbp DNA molecules with a separation quality of ~94%.},
  author       = {Kühnlein, Alexandra},
  keyword      = {Biophysics,microfluidics,DLD,DNA,sorting},
  language     = {eng},
  note         = {Student Paper},
  title        = {Handling of long DNA - applications and polymer physics},
  year         = {2016},
}