Elsevier

Analytical Biochemistry

Volume 179, Issue 1, 15 May 1989, Pages 167-170
Analytical Biochemistry

Large-scale purification of plasmid insert DNA sequences using low-percentage agarose exclusion chromatography

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Abstract

Linear double-stranded DNA fragments ranging from 0.14 to 8.4 kbp have been fractionated on lowpercentage agarose exclusion gels. Both Ultragel A2 (2% agarose) and Bio-Gel A150m (1% agarose) exclude DNA fragments > 900 bp, while the exclusion limit of Bio-Gel A50m (2% agarose) is about 350 bp. All gels result in moderate resolution of DNA fragments smaller than the exclusion limits; we generally observe nearly complete one-step separation of fragments that differ in size by a factor of 2. On the basis of these results, we have used these exclusion gels to routinely purify > 0.4 mg of plasmid insert DNA sequences in one step and over 2.5 mg with a single column, demonstrating that these gel matrices can be ideally suited for repeated rapid large-scale purification of plasmid inserts. In addition, this knowledge allows for a more rational design of plasmids in those cases where large-scale use of the insert DNA is required.

References (11)

  • MaxamA.N. et al.
  • SimpsonR.T. et al.

    Cell

    (1985)
  • MicardD. et al.

    Anal. Biochem.

    (1985)
  • ManiatisT. et al.
  • AusubelF.M. et al.
There are more references available in the full text version of this article.

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This work was supported by NIH Grant GM11719.

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