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MICROCOSM FOR MEASURING SURVIVAL AND CONJUGATION OF GENETICALLY ENGINEERED BACTERIA IN RHIZOSPHERE ENVIRONMENTS
Citation:
Walter, M., L. Porteous, V. Prince, L. Ganio, AND R. Seidler. MICROCOSM FOR MEASURING SURVIVAL AND CONJUGATION OF GENETICALLY ENGINEERED BACTERIA IN RHIZOSPHERE ENVIRONMENTS. U.S. Environmental Protection Agency, Washington, D.C., EPA/600/J-91/141 (NTIS PB91218990).
Description:
A microcosm is described to evaluate and measure bacterial conjugation in the rhizosphere of barley and radish with strains of Pseudomonas cepacia. he purpose was to describe a standard method useful for evaluating the propensity of genetically engineered microorganisms (GEMs) to transfer DNA to recipient bacteria. esults demonstrated the formation of transconjugants from the rhizosphere of each plant 24 h after inoculation. ransconjugant populations peaked at 1.8 X 10 2 colony forming units (CFU)/g root and associated soil in barley and 2.0 X 10 2 CFU/g root and associated soil in radish; they then declined over the next five days of the experiment. o significant differences are found in the survival of transconjugant populations monitored from the two plant species. he microcosm was also used to document the formation of false positive transconjugants, which resulted from donor and recipient P. cepacia mating on the surface of selective agar plates instead of in microcosms. ransconjugants resulting from such plate mating occurred in substantial numbers during the first 5 days of the experiment but declined to undetectable numbers by day 7. he use of nalidixic acid was investigated to determine the magnitude of plate mating. he number of transconjugants detected from radish rhizosphere was reduced by two orders of magnitude by including nalidixic acid in the plating medium; this indicated that 99% of the transconjugants were a result of plate mating.