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Stringent control of transgene expression in Arabidopsis thaliana using the Top10 promoter system
Author(s) -
Love John,
Scott Amie C.,
Thompson William F.
Publication year - 2000
Publication title -
the plant journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.058
H-Index - 269
eISSN - 1365-313X
pISSN - 0960-7412
DOI - 10.1046/j.1365-313x.2000.00706.x
Subject(s) - arabidopsis thaliana , green fluorescent protein , transgene , microbiology and biotechnology , psychological repression , repressor , tetracycline , biology , expression vector , nicotiana tabacum , chemistry , gene expression , gene , mutant , genetics , recombinant dna , antibiotics
Summary We show that the tightly regulated tetracycline‐sensitive Top10 promoter system (Weinmann et al . Plant J. 1994, 5, 559–569) is functional in Arabidopsis thaliana . A pure breeding A. thaliana line (JL‐tTA/8) was generated which expressed a chimeric fusion of the tetracycline repressor and the activation domain of Herpes simplex virus (tTA), from a single transgenic locus. Plants from this line were crossed with transgenics carrying the ER‐targeted green fluorescent protein coding sequence ( mGFP 5 ) under control of the Top10 promoter sequence. Progeny from this cross displayed ER‐targeted GFP fluorescence throughout the plant, indicating that the tTA–Top10 promoter interaction was functional in A. thaliana . GFP expression was repressed by 100 ng ml −1 tetracycline, an order of magnitude lower than the concentration used previously to repress expression in Nicotiana tabacum . Moreover, the level of GFP expression was controlled by varying the concentration of tetracycline in the medium, allowing a titred regulation of transgenic activity that was previously unavailable in A. thaliana . The kinetics of GFP activity were determined following de‐repression of the Top10::mGFP 5 transgene, with a visible ER‐targeted GFP signal appearing from 24 to 48 h after de‐repression.