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Identification and mutational analysis of the promoter for a spinach chloroplast transfer RNA gene.

Authors
  • Gruissem, W
  • Zurawski, G
Type
Published Article
Journal
The EMBO journal
Publication Date
Jul 01, 1985
Volume
4
Issue
7
Pages
1637–1644
Identifiers
PMID: 2992936
Source
Medline
License
Unknown

Abstract

A transcription extract from purified spinach chloroplast was used to test chloroplast DNA sequences for their function as promoter elements. Chloroplast tRNA genes are correctly transcribed in the extract by a soluble RNA polymerase, and precursor molecules are processed into mature tRNAs. Transcription of the spinach chloroplast tRNA2Met gene (trnM2) in vitro requires 5' upstream DNA sequences. Deletion of 5' DNA sequences with exonuclease Bal31 was used to establish the 5' boundary of the promoter region. This boundary is part of a DNA sequence with partial homology to the prokaryotic -35 region. Seventeen base pairs downstream from this sequence a DNA sequence occurs which is homologous to the prokaryotic -10 region. We used synthetic oligonucleotides fused to trnM2 5' deletion mutants to create insertions, deletions and base substitutions in these regions. Internal deletion mutants demonstrated that the -10 promoter element is also required for transcription in vitro. The arrangement of DNA sequences recognised by the chloroplast RNA polymerase resembles the prokaryotic promoter organization.

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