Electrophoretic stretching of DNA adsorbed on supported lipid bilayer in a microcontraction
Date Issued
2011
Date
2011
Author(s)
Chen, Hsin-Wei
Abstract
We perform DNA stretching on supported lipid bilayer (SLB) set on the surface of glass by electric field and electric field gradient. This study is inspired by the recent development of the technology for direct DNA gene mapping. It has received much attention because it enables efficient determination of useful genomic information from DNA. However, the ability to stably and efficiently stretch DNA is the key to the success of this technology.
The proposed method is different from typical DNA stretching techniques because in our case DNA is adsorbed on a lipid bilayer, and therefore DNA stretching happens on 2-dimensional (2D) plane, not in 3-dimensional (3D) space. DNA confined in 2D has a larger equilibrium size and also relaxes slower. In addition, a DNA molecule loses less entropy if it is stretched from 2D configuration relative to from 3D configuration. Therefore, it is expected that stretching DNA on 2D plane will be easier than stretching DNA in 3D space.
Our experiments were performed in a microfluidic channel, and the field gradient was generated by a microcontraction. We find that DNA can be stretched to 70~75% of its contour length. Surprisingly, however, DNA stretching is not caused by the electric field gradient but by other mechanisms. We have observed three different types of DNA stretching. DNA can be hooked by a fixed post, hooked by mobile imprints or tethered to stretch. The formal two have been reported in literatures, while the later is first found in this study. The presence of tethered DNA has been found to be caused by the overhangs of our DNA. We have also studied how the average DNA extension and the probability distribution vary with the electric field strength for different types of DNA stretching. The influence of the charge density of lipid bilayer to DNA stretching has also been investigated.
The cause of the failure to stretch DNA by electric field gradient is still unclear. However, DNA has often been observed to rapidly retract once it is stretched over a certain degree. We believe this is strongly related to the breakdown of our expectation, and more investigation is required to resolve this mystery.
Subjects
DNA stretching
supported lipid bilayer
microcontraction
electrophoresis
Type
thesis
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