R‐Loop Mapping of Calf‐Thymus RNA Polymerase II. Transcripts in vitro on Restriction Fragments of Adenovirus 2 DNA
1982
Spindler, Ernst | Stein, Hans
Nascent RNA, synthesized by calf thymus RNA polymerase II on the restriction endonuclease EcoRI fragment A and BamHI fragment B of adenovirus 2 DNA, was rehybridized to the template strand under conditions allowing transcription R‐loop formation. Hybrids, visualized by electron microscopy, were found in looped and in branched configurations, the former being abundant, with an average loop number of roughly four per EcoRI fragment A and three per BamHI fragment B. Frequency distributions of transcription R‐loops, synthesized on identical restriction fragments for different periods of time, showed similar patterns, thus pointing to a nonrandom type of transcription. A comparison of the frequency distribution of R‐loops seen on the two overlapping restriction fragments reveals reasonable coincidence in the loop pattern in one out of the four possible fragment orientations, permitting correlation of the transcription R‐loop pattern with the physical map of the adenovirus 2 DNA. Comparison of start sites tentatively mapped in vitro with the known promoters for RNA polymerase II in vivo suggests that a major transcription site in vitro is located in close proximity to, or identical with, two promoters in vivo at 16.1 and 16.5 map units
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