Isolation of growth-phase-specific promoters from cultured tobacco cells

Strong promoters are required under several culture conditions for effective transgene expression in tobacco BY2 cells. We have isolated the promoter fragments of 4 genes exhibiting high homology to those of Arabidopsis thaliana 108C1T7 (unknown function) and F1-ATPase-delta, alcohol dehydrogenase and pectin esterase genes from a genomic DNA library of BY2 cells. Two of the four genes were strongly expressed during every phase of growth of BY2 cells, and the other two were expressed only during the stationary phase. Each of the promoter fragments was ligated to the GUS reporter gene and introduced into the chromosome of BY2 cells by Agrobacterium-mediated transformation. Growth-phase-dependent expression of the GUS gene was reproduced under the control of all 4 promoters observed with the original genes. Significantly higher expression was observed under the control of Nt108p during every phase of cell growth and under the control of NtADHp and NtPESp during the stationary phase than that under the control of the CaMV35S promoter.