Vacuum blotting: a simple method for transferring DNA from sequencing gels to nylon membranes

David S. Gross, Kerry W. Collins, Efrem M. Hernandez, William T. Garrard

Research output: Contribution to journalArticle

10 Scopus citations

Abstract

We describe a vacuum blotting procedure for transferring DNA fragments from conventional polyacrylamide sequencing gels to nylon membranes. The method employs a combination of vacuum-assisted diffusion (effected by a standard gel drier) and an osmotic gradient (effected by over- and underlying filters presoaked in ammonium acetate). Fragments up to 310 nucleotides in length transfer at 40-60% efficiency within 90 min. When combined with indirect end-labelling, the method allows genomic sequencing of a single-copy gene of Saccharomyces cerevisiae employing as little as 5 μg DNA per lane.

Original languageEnglish (US)
Pages (from-to)347-356
Number of pages10
JournalGene
Volume74
Issue number2
DOIs
StatePublished - Dec 30 1988

Keywords

  • Genomic sequencing
  • RNA hybridization probes
  • in vivo footprinting
  • indirect end-labelling

ASJC Scopus subject areas

  • Genetics

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