Preparative-scale isolation of multiple genes by agarose-based continuous elution electrophoresis

Journal Article (Journal Article)

Background. Techniques for the isolation of genes from restriction enzyme size fractionated genomic DNA are traditionally accomplished by procedures that are both time and labor intensive. Therefore, we developed a simple and efficient procedure for the isolation of genes as individual DNA fragment lengths ranging in size from 1 kb to > 18 kb. Methods. Using a commercial continuous elution protein electrophoresis apparatus and incorporating a non-toxic agarose matrix, preparative-scale amounts (300 μg) of enzyme-restricted genomic DNA was size fractionated by continuous elution electrophoresis, capturing the target DNA sequence of interest. Fractions corresponding to unique fragment length ranges were then screened for individual genes by dot-blot analysis. Results. We have isolated two genes (PGK1; a single copy housekeeping gene, and p53 (exons 3-11); a tumor suppressor gene) from one preparative run, whose DNA fragment lengths elute at 1 kb and 7.5 kb respectively. Although we have performed separation and isolation of DNA fragment lengths from 1 kb to > 18 kb with high recovery yields, different fragment lengths can be isolated by varying the gel concentration in the preparative column. Conclusions. Advantages of this technique are that it is an easy and efficient means of large-scale isolation of DNA fragment lengths, with the added advantage of isolating multiple genes from a single preparative run. Genomic DNA fragment length purification by agarose matrix continuous elution electrophoresis should prove useful in research areas which require gene isolation, such as genomics and molecular biology.

Duke Authors

Cited Authors

  • Rodriguez, H; Akman, SA

Published Date

  • December 1, 1998

Published In

Volume / Issue

  • 6 / 3

Start / End Page

  • 150 - 155

International Standard Serial Number (ISSN)

  • 1122-8652

Citation Source

  • Scopus