HEAT-INDUCIBLE EXPRESSION SYSTEM FOR A FOREIGN GENE IN CULTURED TOBACCO CELLS USING THE HSP18.2 PROMOTER OF ARABIDOPSIS-THALIANA

A system for the controlled expression of a foreign gene in cultured t obacco cells (Nicotiana tabacum, BY2) by temperature shift was constru cted. A 925-base-pair (bp) DNA fragment containing the 5'-flanking reg ion of a low-molecular-mass heat-shock protein gene (HSP18.2) of Arabi dopsis thaliana was inserted upstream of the beta-glucuronidase report er gene (GUS). The resulting HSP18.2-GUS construct was introduced into BY2 cells by electroporation or Agrobacterium-mediated transformation . Transient expression of the HSP18.2 promoter in protoplasts was very low regardless of the heat shock. Although expression of the HSP18.2- GUS chimeric gene in the stable transformants of BY2 was hardly detect ed in culture at 25 degrees C, the expression increased rapidly on the transcriptional level when the incubation temperature was shifted to 35-37 degrees C. The optimal temperature for heat-shock induction was 37 degrees C. After a 2-h incubation at 37 degrees C, GUS activity was about 1000-fold greater than that before heat shock. The amount of GU S mRNA was maximum 2 h after heat shock, and then decreased gradually.