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Nucleic Acids Research, 1979, Vol. 7, No. 7 1851-1868
© 1979

Articles

Gel electrophoretic separation of transcription complexes: an assay for RNA polymerase selectivity and a method for promoter mapping

Barry K. Chelm and E.Peter Geiduschek

Department of Biology, University of California at San Diego La Jolla, CA 92093, USA

Received September 5, 1979. We describe a method for analyzing ternary transcription complexes, of RNA polymerase, DNA and nascent RNA,chains, by agarose gel electrophoresis. fchen the RNA of such complexes is P-labelled, a simple comparison of the DNA fluorogram with an autoradiogram identifies transcriptionally active DNA molecules and restriction fragments in any mixture. Two limitations on the method are described: 1) retardation during electrophoresis of polymerase-DNA complexes relative to their conjugate bare DNA fragments; 2) failure of very large ternary complexes to enter gels.

The following potential applications of the method are surveyed: transcription unit (elongation) mapping, separation of RNA molecules in a mixture of transcripts, dinucleotide primer mapping and identification of preferred template conformations.


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