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March 11, 1983Nucleic Acids Research11,634 citationsOpen Access

Accurate transcription initiation by RNA polymerase II in a soluble extract from isolated mammalian nuclei

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JDJohn David DignamUniversity of ToledoRLRussell M. LebovitzAmprion (United States)RRR G RoederRockefeller University

Key Points

  • The aim is to develop a procedure for producing nuclear extracts that enable accurate transcription initiation in vitro.
  • Prepared extracts from nuclei of human tissue culture cells.
  • Optimized conditions for maximum activity using adenovirus 2 major late promoter.
  • Assayed transcription initiation from class II and class III promoters.
  • Achieved accurate transcription initiation from class II and III promoters.
  • Extract directed transcription initiation from adenovirus and cellular promoters.
  • Demonstrated compatibility with tRNA and Ad 2 VA promoters.

Abstract

We have developed a procedure for preparing extracts from nuclei of human tissue culture cells that directs accurate transcription initiation in vitro from class II promoters. Conditions of extraction and assay have been optimized for maximum activity using the major late promoter of adenovirus 2. The extract also directs accurate transcription initiation from other adenovirus promoters and cellular promoters. The extract also directs accurate transcription initiation from class III promoters (tRNA and Ad 2 VA).

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Cite This Study

Dignam et al. (1983) studied this question.

synapsesocial.com/papers/69df0fc9d85e58e37b7a114dhttps://doi.org/10.1093/nar/11.5.1475
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