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DNA Isolation

DNA Isolation

DNA isolation protocols can be fast and straightforward or complex and time-consuming depending on the type and quality of DNA required for further processing. For example, plasmid DNA extraction is a bit more complicated than genomic DNA extraction because of the need for an appropriate lysis method to separate plasmid DNA from gDNA during isolation. However, for specific applications, such as long-range DNA sequencing that require a good yield of high- quality DNA samples, we need to follow...

DNA Isolation

DNA Isolation

DNA from cells is required for many biotechnology and research applications, such as molecular cloning. To remove and purify DNA from cells, researchers use various methods of DNA extraction. While the specifics of different protocols may vary, some general concepts underlie the process of DNA extraction.

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Affinity Chromatography

Affinity chromatography is a powerful technique extensively utilized for separating and purifying specific biomolecules from complex mixtures. It capitalizes on the highly selective binding between an analyte and its counterpart, such as antibody-antigen interactions. The counterpart is immobilized on the stationary phase, forming an affinity column. The stationary phase typically consists of solid support, such as agarose or porous glass beads, immobilizing the affinity ligand. The mobile...

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Related Experiment Video

Updated: Jun 17, 2026

Streamlined Purification of Plasmid DNA From Prokaryotic Cultures

Streamlined Purification of Plasmid DNA From Prokaryotic Cultures

Published on: January 5, 2011

Dual affinity method for plasmid DNA purification in aqueous two-phase systems.

H S C Barbosa¹, A V Hine, S Brocchini

¹Center of Chemistry, University of Minho, Campus de Gualtar, 4710-057 Braga, Portugal.

Journal of Chromatography. A

|January 20, 2010

Summary

This study introduces a dual affinity system for plasmid DNA extraction using a fusion protein and a functionalized polymer in aqueous two-phase systems. The method shows promise for efficient DNA purification from bacterial lysates.

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Last Updated: Jun 17, 2026

Streamlined Purification of Plasmid DNA From Prokaryotic Cultures

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Published on: January 5, 2011

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Purifying Plasmid DNA from Bacterial Colonies Using the Qiagen Miniprep Kit

Purifying Plasmid DNA from Bacterial Colonies Using the Qiagen Miniprep Kit

Published on: July 29, 2007

Area of Science:

Biotechnology
Molecular Biology
Biochemistry

Background:

Plasmid DNA extraction is crucial for molecular biology applications.
Traditional methods can be time-consuming and may lead to contamination.
Aqueous two-phase systems (ATPS) offer a scalable alternative for bioseparation.

Purpose of the Study:

To evaluate a dual affinity system for pUC19 plasmid DNA extraction.
To utilize poly(ethylene glycol)/dextran (PEG/DEX) ATPS combined with a fusion protein and a functionalized polymer.
To assess the efficiency and purity of plasmid DNA extracted from bacterial cell lysate.

Main Methods:

A dual affinity system involving LacI-His6-GFP fusion protein and PEG-IDA-Cu(II) conjugate was employed.
Plasmid DNA extraction was performed within PEG/DEX aqueous two-phase systems.
Agarose gel electrophoresis was used to evaluate DNA purity and contamination by RNA or genomic DNA.

Main Results:

Over 72% of plasmid DNA partitioned to the PEG phase in the first extraction stage with high purity.
Elution of plasmid DNA from the fusion protein complex was challenging in the second stage.
A maximum recovery of approximately 27% of pUC19 plasmid DNA was achieved, yielding 7.4 μg/mL of lysate.

Conclusions:

The dual affinity system in PEG/DEX ATPS demonstrates potential for initial plasmid DNA enrichment.
Optimization of the elution step is necessary for improved overall recovery.
This approach offers a novel strategy for plasmid DNA purification, particularly for large-scale applications.