Generation of Conditional Mutants in Higher Eukaryotes by Switching between the Expression of Two Genes

A regulatory system for the in-depth study of gene functions in higher eukaryotic cells has been developed. It is based on the tetracycline-controlled transactivators and reverse tTA, which were remodeled to discriminate efficiently between two different promoters. The system permits one to control...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 1999-02, Vol.96 (3), p.1013-1018
Main Authors: Baron, Udo, Schnappinger, Dirk, Helbl, Vera, Gossen, Manfred, Hillen, Wolfgang, Bujard, Hermann
Format: Article
Language:English
Subjects:
Bacterial Proteins - genetics
Biological Sciences
Cell lines
Dimerization
DNA
DNA-Binding Proteins - metabolism
Doxorubicin - toxicity
Gene Expression Regulation, Bacterial - drug effects
Genes
Genes, Switch
Genetics
HeLa Cells
Humans
Mutation
Phenotype
Phenotypes
Plasmids
Promoter Regions, Genetic - drug effects
Proteins
Recombinant Fusion Proteins - metabolism
Tetracycline - pharmacology
Trans-Activators - genetics
Trans-Activators - metabolism
Transactivators
Transcriptional activation
Transcriptional Activation - drug effects
Transfection
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Summary:A regulatory system for the in-depth study of gene functions in higher eukaryotic cells has been developed. It is based on the tetracycline-controlled transactivators and reverse tTA, which were remodeled to discriminate efficiently between two different promoters. The system permits one to control reversibly the activity of two genes, or two alleles of a gene, in a mutually exclusive way, and also allows one to abrogate the activities of both. This dual regulatory circuit, which can be operated by a single effector substance such as doxycycline, overcomes limitations of conventional genetic approaches. The conditional mutants that can now be generated will be useful for the study of gene function in vitro and in vivo. In addition, the system may be of value for a variety of practical applications, including gene therapy.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.96.3.1013