Regeneration of commercial nucleic acid extraction columns without the risk of carryover contamination

Siddappa Nagadenahalli Byrareddy, Appukuttan Avinash, Mohanram Venkatramanan, Udaykumar Ranga

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

Nucleic acid extraction is a basic requirement in a molecular biology laboratory. In terms of purity and yield, commercial nucleic acid extraction columns are superior; however, they are expensive. We report here an efficient strategy to regenerate diverse commercial columns for several rounds without altering the binding capacity of the columns or changing the properties of the nucleic acids purified. Plasmids purified with regenerated columns were functionally identical in super-coiled nature, restriction analysis, expression of the encoded reporter genes, or amplification of the viral RNA in real-time PCR. To ensure that the regenerated columns were free of the residual DNA, we used two different plasmids with different drug-resistance markers. By colony plating and PCR amplification of the encoded genes, we show that the regeneration process is absolute. Using radiolabeled DNA, we demonstrate that DNA exposed to the regeneration reagent is fragmented to molecular weight below 36 bp. Our data collectively prove regeneration of the commercial columns without the concern of carryover contamination. A procedure to permit safe and efficient regeneration of the commercial columns is not only of great advantage to extend the lifetime of these columns but also makes them commercially more affordable, especially in a resource-poor setting.

Original languageEnglish (US)
Pages (from-to)186-192
Number of pages7
JournalBioTechniques
Volume42
Issue number2
DOIs
StatePublished - Feb 1 2007

Fingerprint

Nucleic Acids
Regeneration
Contamination
Amplification
Gene Amplification
DNA
Plasmids
Genes
Molecular biology
Viral RNA
Plating
Molecular weight
Reporter Genes
Drug Resistance
Real-Time Polymerase Chain Reaction
Molecular Biology
Molecular Weight
Pharmaceutical Preparations
Polymerase Chain Reaction

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

Regeneration of commercial nucleic acid extraction columns without the risk of carryover contamination. / Byrareddy, Siddappa Nagadenahalli; Avinash, Appukuttan; Venkatramanan, Mohanram; Ranga, Udaykumar.

In: BioTechniques, Vol. 42, No. 2, 01.02.2007, p. 186-192.

Research output: Contribution to journalArticle

Byrareddy, Siddappa Nagadenahalli ; Avinash, Appukuttan ; Venkatramanan, Mohanram ; Ranga, Udaykumar. / Regeneration of commercial nucleic acid extraction columns without the risk of carryover contamination. In: BioTechniques. 2007 ; Vol. 42, No. 2. pp. 186-192.
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