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Iterative Optimization of DNA Duplexes for Crystallization of SeqA-DNA Complexes
Published on: November 1, 2012
Poly(acrylamide-co-alkylacrylamides) for electrophoretic DNA purification in microchannels
Thomas N Chiesl1, Wei Shi, Annelise E Barron
1Department of Chemical and Biological Engineering, Northwestern University, Evanston, IL 60208, USA.
Analytical Chemistry
|February 1, 2005
Summary
New water-soluble copolymers selectively remove proteins from DNA using microchannel electrophoresis. Hydrophobic N-alkylacrylamide subunits enable protein adsorption, crucial for genetic analysis applications.
Area of Science:
- Polymer Chemistry
- Analytical Chemistry
- Biotechnology
Background:
- Selective separation of proteins from DNA is essential for genetic analysis.
- Microchannel electrophoresis requires advanced matrix materials for efficient separation.
- Hydrophobic interactions are a key mechanism for molecular adsorption.
Purpose of the Study:
- To synthesize and characterize water-soluble block copolymers for selective protein removal from DNA.
- To investigate the effect of hydrophobic N-alkylacrylamide subunits on protein adsorption.
- To evaluate the performance of these copolymers in microchannel electrophoresis for genetic analysis.
Main Methods:
- Synthesis of N-alkylacrylamide comonomers with varying alkyl chain lengths (C4, C6, C8, C6-C6).
- Aqueous "micellar" polymerization technique to create block copolymers.
- Characterization using 1H NMR, GPC-MALS, and capillary electrophoresis.
- On-chip microchannel electrophoresis experiments.
Main Results:
- Copolymers with octylacrylamide and dihexylacrylamide subunits demonstrated significant protein adsorption.
- Butylacrylamide copolymers showed minimal protein adsorption.
- All synthesized copolymer matrices allowed dsDNA passage and exhibited DNA sieving properties.
- Successful demonstration of on-chip protein adsorption using the developed matrices.
Conclusions:
- Water-soluble block copolymers incorporating hydrophobic N-alkylacrylamide subunits effectively adsorb proteins.
- These novel polymer matrices show promise for selective protein removal in microfluidic devices.
- The developed materials advance the goals of micro Total Analysis Systems (µ-TAS) for genetic analysis.
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Gel electrophoresis is a method that separates biological macromolecules like nucleic acids or proteins by forcing them to pass through a gel matrix under an electric field.
A variation of gel electrophoresis, termed polyacrylamide gel electrophoresis (PAGE), is commonly used for separating proteins according to their molecular size by passing them through a polyacrylamide gel. Because of the varying charges associated with amino acid side chains, PAGE can be used to separate intact proteins...
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Two-dimensional gel electrophoresis is a high-resolution protein separation method first introduced by O' Farrell and Klose in 1975. This method involves protein separation by two dimensions, mass and charge, making it more accurate than one-dimensional gel electrophoresis.
The first dimension separation uses the isoelectric focusing or IEF technique performed on immobilized pH gradient (IPG) strips that separate proteins according to their isoelectric points.
Biological samples, such as cells...
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