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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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Related Experiment Video

Updated: Jun 3, 2026

Streamlined Purification of Plasmid DNA From Prokaryotic Cultures

Streamlined Purification of Plasmid DNA From Prokaryotic Cultures

Published on: January 5, 2011

Plasmid DNA isolation (sheared lysate method).

J W Dale¹, P J Greenaway

¹Department of Microbiology, University of Surrey, Guildford, Surrey.

Methods in Molecular Biology (Clifton, N.J.)

|March 5, 2011

Summary

This summary is machine-generated.

The cleared lysate method struggles with large plasmid DNA isolation. Cesium chloride-ethidium bromide gradients offer an effective alternative for recovering high molecular weight plasmids from cell lysates.

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Last Updated: Jun 3, 2026

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Published on: July 29, 2007

Area of Science:

Molecular Biology
Biochemistry

Background:

The cleared lysate method is commonly used for plasmid DNA isolation.
This method is often ineffective for plasmids exceeding 20 kb.
Poor recovery of large plasmids is attributed to their removal during the clearing spin step.

Purpose of the Study:

To address the limitations of the cleared lysate method for large plasmid isolation.
To present an alternative, more effective procedure for recovering high molecular weight plasmid DNA.

Main Methods:

Utilizing a cesium chloride-ethidium bromide gradient.
Loading the complete cell lysate directly onto the gradient.
Separating plasmid DNA from chromosomal DNA and other cellular components via density gradient centrifugation.

Main Results:

The cesium chloride-ethidium bromide gradient method effectively separates plasmid DNA.
This alternative procedure overcomes the limitations of the cleared lysate method for large plasmids.
Improved recovery of high molecular weight plasmid DNA is achieved.

Conclusions:

Cesium chloride-ethidium bromide density gradient centrifugation is a superior method for isolating large plasmids (>20 kb).
This technique provides better recovery of plasmid DNA compared to the cleared lysate approach.
The method effectively separates plasmid DNA from genomic DNA and other cellular contaminants.