DNA separation by silica adsorption is a DNA separation method based on DNA molecules that bind to the silica surface in the presence of a particular salt and under certain pH conditions.
Video DNA separation by silica adsorption
Significance
Conventional methods for DNA extraction, such as ethanol deposition or preparation using commercial purification kits, can not be integrated into microchips because they require several direct processing steps. In addition, they also require large equipment and high volume of reagents and samples. DNA extraction on microchips provides a fast, effective, and effective cost for high throughput filtering, which also has a very small footprint. This new method has useful applications for biosensors, labs on chip devices, and other new technologies that require fast, high quality DNA at minimal cost.
Maps DNA separation by silica adsorption
Operation
In actual operation, the sample (this may be anything from the purified cell to the tissue specimen) is placed into the chip and lysed. The resulting mixture of proteins, DNA, phospholipids, etc., is then run through channels in which the DNA is adsorbed by the silica surface in the presence of solutions with high ionic strength. The highest DNA adsorption efficiency occurs in the presence of buffer solutions with pH at or below the pKa of the surface silanol group.
Although this mechanism is not fully understood, one possible explanation involves the reduction of the negative surface load of silica due to the high ionic strength of the buffer. This decrease in surface charge causes a decrease in electrostatic repulsion between negatively charged DNA and negatively charged silica. Meanwhile, buffers also reduce water activity by formatting hydrated ions. This leads to the surface of the silica and the DNA becomes dehydrated. These conditions lead to a very favorable situation for DNA to be absorbed onto the surface of silica.
A further explanation of how DNA binds silica is based on the action of guanidium HCl (GuHCl), which acts as chaotrop. A chaotrope alters the biomolecule by interfering with the surrounding hydration shell. This allows positively charged ions to form salt bridges between negatively charged silica and the backbone of negatively charged DNA in high salt concentrations. DNA can then be washed with high salt and ethanol, and ultimately elute with low salt.
After the DNA is adsorbed onto the silica surface, all other molecules pass through the column. Most likely, these molecules are sent to the waste part of the chip, which can then be closed using a wakeful channel or pressure or controlled voltage chamber. The DNA is then washed to remove excess waste particles from the sample and then eluted from the channel using an elution buffer for further downstream processing.
The following solutions have been proposed and validated for use in this binding DNA process: the GuHCl-based loading buffer; Channel Wash: 80% isopropanol; DNA Elution: TE at pH 8.4.
Silicon microstructure DNA surface
Methods using silica beads and silica resins have been created that can isolate DNA molecules for subsequent PCR amplification. However, this method has an associated problem. First, the beads and resins vary greatly depending on how well they are packaged and thus difficult to reproduce. Each micro channel loading can generate different packing numbers and thus change the amount of adsorbed DNA to the channel. Furthermore, this method produces a two-step process.
The silica structure is a much more effective method of packaging because it is etched into the canal during its manufacture and thus is the result of a one step process through soft lithography. The silica structure is easier to use in very parallel designs than beads or resins.
See also
- Biochip
- Boom Method
- Downstream processing
- Microelectromechanical system
- Microfluidics
- Rotate purification of nucleic acids by column
References
- Cady, et al. Purification of nucleic acids using a microfabricated silicon structure. Biosensor and Bioelectronics, 19, 59-66 (2003).
- K. A. Melzak, C. S. Sherwood, R. F. B. Turner, C. A. Haynes. Driving Forces to Adsorption of DNA into Silica in Perchlorate Solution. Journal of Colloidal Science and Interface, 181, 635-644 (1996).
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- Wolfe, et al. Toward a microchip-based solid phase extraction method for nucleic acid isolation. Electrophoresis, 23, 727-733 (2002).
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