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Size-based separation of supercoiled plasmid DNA using ultrafiltration
Ying Li1, Neil Butler1, Andrew L Zydney1
1Department of Chemical Engineering, The Pennsylvania State University, University Park, PA 16802, United States.
Journal of Colloid and Interface Science
|April 3, 2016
Summary
This study reveals that DNA size significantly impacts plasmid transmission through ultrafiltration membranes, enabling size-based separations. Supercoiled plasmid branching allows for selective separation based on DNA length.
Area of Science:
- Biotechnology
- Molecular Biology
- Separation Science
Background:
- Previous studies indicated no DNA size effect on plasmid transmission through ultrafiltration membranes, aligning with flexible polymer behavior.
- Supercoiled plasmids exhibit a branched structure, with branching complexity correlating to DNA length.
- This structural difference presents an opportunity for size-based plasmid separations.
Purpose of the Study:
- To investigate the influence of plasmid size and supercoiling on transmission through ultrafiltration membranes.
- To explore the potential for size-based separation of plasmids using ultrafiltration technology.
Main Methods:
- Utilized 3.0, 9.8, and 16.8 kbp plasmids (both supercoiled and linear isoforms).
- Employed cellulosic and polyethersulfone ultrafiltration membranes with varying molecular weight cutoffs.
- Analyzed plasmid transmission under different filtrate flux conditions.
Main Results:
- Plasmid transmission increased with filtrate flux due to flow-induced plasmid elongation.
- Flux dependence varied among plasmids due to differing degrees of supercoiled DNA branching.
- Achieved significant selectivity for separating 3.0 and 16.8 kbp supercoiled plasmids.
Conclusions:
- Plasmid branching significantly influences transmission through ultrafiltration membranes, contrary to previous findings for linear DNA.
- Ultrafiltration can be optimized for size-based separation of supercoiled plasmids.
- This research opens avenues for advanced plasmid purification techniques.
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