Efficiency of physical (light) or chemical (ABA, tetracycline, CuSO4 or 2-CBSU)-stimulus-dependent gus gene expression in tobacco cell suspensions.
In this study, the efficiency of inducible promoters to switch on gene expression in the presence of inducer or to switch it off in its absence was evaluated in tobacco cell suspensions transformed with the gus gene coding sequence. Either plant (pats1A, pSalT, pIn2-2) or microbial (pMre, pTet) inducible promoters were used to drive gus expression. The inducers were light, abscisic acid, 2-CBSU, CuSO4, tetracycline, respectively. For each construct (inducible promoter-gus coding sequence), the optimal induction conditions were determined (inducer concentration, induction time, and age of cells in culture cycle before induction). The efficiency of the inducible promoter was then evaluated under optimal induction conditions. GUS-expression levels obtained under non-inducing and inducing conditions were systematically compared. Thirty or forty percent of the clones transformed with the pSalT-gus or pTet-gus construct, respectively, showed high induction rates (>1000) and GUS activities of the same order as those obtained with a constitutive system. However, basal GUS levels were always high for the pTet-gus cell lines. Seventy or eighty-five percent of the cell lines transformed with the pMre-gus or pln2-2-gus construct, respectively, had induction rates of 1.5 to 1000. The pats1A-gus construct gave very low induction rates-55% of cell lines had induction rates less than 1.5. Only the pSalt-gus construct gave both the highest induction rates and basal GUS-levels equivalent to the endogenous GUS background.
Boetti
H
H
Laboratoire de Technologie Enzymatique, UPRES A 6022 CNRS, Université de Technologie de Compiègne, BP 20529, 60205 Compiegne Cedex, France. boetti@u-cergy.fr
Chevalier
L
L
Denmat
L A
LA
Thomas
D
D
Thomasset
B
B
D000040:Abscisic Acid / Q000494:pharmacology; D016960:Agrobacterium tumefaciens / Q000235:genetics; D002478:Cells, Cultured; D003001:Cloning, Molecular; D019327:Copper Sulfate / Q000494:pharmacology; D004926:Escherichia coli / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects* / Q000528:radiation effects*; D005966:Glucuronidase / Q000235:genetics*; D008027:Light; D010947:Plants, Toxic; D011401:Promoter Regions, Genetic; D013449:Sulfonamides / Q000494:pharmacology; D013752:Tetracycline / Q000494:pharmacology; D014026:Nicotiana / Q000166:cytology / Q000235:genetics*; D000096926:Benzenesulfonamides
D000040:Abscisic Acid; D019327:Copper Sulfate; D005966:Glucuronidase; D013449:Sulfonamides; D013752:Tetracycline; C071356:N-(aminocarbonyl)-2-chlorobenzenesulfonamide; D000096926:Benzenesulfonamides