Studies on the photosynthesis-inhibiting herbicide resistance of photoautotrophically cultured tobacco cells

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The effects of herbicides with different primary modes of action were examined on the growth of photoautotrophic, photomixotrophic, heterotrophic cultures of tobacco cells. These responses were compared with those of tobacco seedlings to the same herbicides. Herbicides , which primarily inhibit or disturb photosynthetic processes, suppressed the growth of photoautotrophic cells most strongly, as compared to photomixotrophic and heterotrophic cells (atrazine, diuron, paraquat). Herbicides having a primary mode of action other than the inhibition of photosynthetic processes, suppressed the growth of all type of cultured cells at similar concentration (2,4-D, diphenamid, glyphosate, dinoseb, sodium chlorate, bialaphos, DTP). Photoautotrophic cells were the most sensitive cultured cell lines to all kinds of herbicides except sodium chlorate. Furthermore, photoautotrophic cells responded to most of the herbicides as did the seedlings, with the exception of glyphosate and diphenamid. It was concluded that photoautotrophically cultured cells would be the most suitable system to study the effects of herbicides in vitro.Based on the above information, a variant cell line showing resistance to the photosynthesis-inhibiting hervicide atrazine was selected from cultured photoautotrophic tobacco cells by repeated exposure to toxic levels of the herbicide. The resistant cell line was shown to have a point mutation in the chloroplast psbA gene. Furthermore, this mutation subsequently was stavle in the absence of continued selection pressure. This demonstrate that photoautotrophic cell culture can be used for genetic engineering of the chloroplast genome.