Herbicide tolerance in transgenic plants expressing bacterial detoxification genes. The case of bromoxynil

Introduction of detoxification genes into crops provides the opportunity to enhance the selectivity of herbicides. In this paper we present details of the use of a bacterial nitrilase in conferring resistance to the oxynil (bromoxynil and ioxynil) herbicides. A gene coding for a specific nitrilase has been cloned from a strain of Klebsiella ozaenae isolated from bromoxynil treated soil. The coding region of this gene (termed oxy) has been put under the control of either the CaMV 35S promoter or the promoter of a RuBisCO small subunit gene. These chimaeric genes have been introduced into tobacco (PBD6) and oilseed rape (Westar) using Agrobacterium transformation with only the "oxy" gene as a selectable marker. Non-transformed plants of both species are susceptible at 100-150g oxynils/ha however, the presence of the "oxy" gene allows transformed plants to tolerate greater than 4,800 g a.i./ha oxynils under field conditions, which is more than 10 times the use rate of these herbicides in crops. The oxynils are inhibitors of photosynthetic electron transport and are also uncouplers of photophosphorylation and their herbicidal activity can be monitored in treated plants using in vivo fluorescence techniques. Fluorescence spectra of transformed oilseed rape treated with 3,000 g a.i./ha bromoxynil closely resembled those of untreated controls with only a small initial increase in fluorescence 10-15 min after spraying which returned to control values within Ih. A close correlation was evident between oxynil degradation and leaf photosynthetic inhibition. Studies were carried out using radio-tie analysis of plants treated with C-14 bromoxynil. Bromoxynil was rapidly degraded in transformed plants to the corresponding non-phytotoxic benzoic acid. Ninety-three percent of the absorbed bromoxynil was degraded in treated leaves within 2h of treatment with a dose equivalent of 3,000 g a.i./ha. Twenty-four hours after treatment 7-8% of the absorbed C-14 activity was translocated from treated leaves of which 94% was benzoic acid and parent bromoxynil was not detected. In contrast, no degradation of bromoxynil was observed in untransformed plants 24h after treatment.