Plant Molecular Biology

, Volume 16, Issue 1, pp 1–10

Gene structure and expression of a tobacco endochitinase gene in suspension-cultured tobacco cells

  • Yuji Fukuda
  • Masaru Ohme
  • Hideaki Shinshi
Article

DOI: 10.1007/BF00017912

Cite this article as:
Fukuda, Y., Ohme, M. & Shinshi, H. Plant Mol Biol (1991) 16: 1. doi:10.1007/BF00017912

Abstract

We have isolated and characterized the genomic clone λCHN50 corresponding to tobacco basic endochitinase (E.C.3.2.1.14). DNA sequence and blotting analysis reveal that the coding sequence of the gene present on λCHN50 is identical to that of the cDNA clone pCHN50 and, moreover, the CHN50 gene has its origin in the progenitor of tobacco, Nicotiana sylvestris. Tobacco basic chitinases are encoded by a small gene family that consists of at least two members, the CHN50 gene and a closely related CHN17 gene which was characterized previously. By northern blot analysis, it is shown that the CHN50 gene is highly expressed in suspension-cultured tobacco cells and the mRNA accumulates at late logarithmic growth phase. To identify cis-DNA elements involved in the expression of the CHN50 gene in suspensioncultured cells, the chimeric gene consisting of 1.1 kb CHN50 5′ upstream region fused to the coding sequence of β-glucuronidase (GUS) was introduced by electroporation into protoplasts isolated from suspension-cultured tobacco cells. Transient GUS activity was found to be dependent on the growth phase of the cultured cells, from which protoplasts had been prepared. Functional analysis of 5′ deletions suggests that the distal region between -788 and -345 contains sequences that potentiate the high-level expression in tobacco protoplasts and the region (-68 to -47) proximal to the TATA box functions as a putative silencer.

Key words

chimeric genechitinase genegene familypromoter regiontransient expression

Copyright information

© Kluwer Academic Publishers 1991

Authors and Affiliations

  • Yuji Fukuda
    • 1
  • Masaru Ohme
    • 1
  • Hideaki Shinshi
    • 1
  1. 1.Fermentation Research InstituteAgency of Industrial Science and TechnologyTsukuba, IbarakiJapan