Denaturation map of bacteriophage T7 DNA and direction of DNA transcription

Abstract

After partial denaturation of bacteriophage T7 DNA by heat or alkali, electron microscopy showed preferential strand separation at A + T-rich sites which map asymmetrically at the following fractional lengths: 0.01, 0.16, 0.21, 0.26, 0.33, 0.42, 0.46, 0.57, 0.63, 0.66, 0.86, 0.91 and 0.99. This map physically distinguishes between the two molecular ends of T7 DNA. In vitro synthesis of RNA by Escherichia coli RNA-polymerase with T7 DNA as a template and subsequent partial heat denaturation of the complexed DNA permitted simultaneous localization of RNA-synthesis sites and of strand-separation sites on individual molecules. Since the enzyme is known to initiate close to the genetically defined left-end of T7 DNA, it was found that the fractional length 0 of the denaturation map is equivalent to the left-end of the genetic map and that at least 90% of RNA synthesis is initiated close to the left-end of T7 DNA.