Snapshot blotting: transfer of nucleic acids and nucleoprotein complexes from electrophoresis gels to grids for electron microscopy

Proc Natl Acad Sci U S A. 1994 Jul 19;91(15):6870-4. doi: 10.1073/pnas.91.15.6870.

Abstract

We present a technique, "snapshot blotting," for the electrophoretic transfer of nucleic acids and nucleoprotein complexes in gel electrophoresis bands onto highly stable carbon film-coated grids for imaging by electron microscopy. The method permits structural analysis of macromolecular species that have been resolved by a gel mobility-shift assay. To demonstrate the efficiency and integrity of the transfer process for a multiprotein-DNA assembly, we have imaged various species of a prokaryotic transcription complex, using the cleavage-defective EcoRI(Q111) protein as an orientation marker and as a blockade of transcription elongation. Snapshot blotting should be of great utility in the structural characterization of nucleic acids and protein-nucleic acid interactions.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • DNA / chemistry
  • DNA / ultrastructure
  • Deoxyribonuclease EcoRI / ultrastructure
  • Electrophoresis, Agar Gel / methods*
  • Microscopy, Electron
  • Nucleic Acids / chemistry
  • Nucleic Acids / ultrastructure*
  • Nucleoproteins / chemistry
  • Nucleoproteins / ultrastructure*
  • Transcription, Genetic

Substances

  • Nucleic Acids
  • Nucleoproteins
  • DNA
  • Deoxyribonuclease EcoRI