Cannabis sativa L. is grown for medicinal and recreational uses in Canada. Factors affecting tissue culture response and genetic transformation using Agrobacterium were investigated. Variables including genotype, explant source, light intensity, explant preparation, Agrobacterium strain, and resistance marker selection influenced the ability to obtain transformed callus, which occurred at a maximum frequency of 65% via RT-PCR. Genetic transformation was confirmed by PCR and RT-PCR using primers to confirm the presence of the bialaphos (bar) selection marker as well as the Arabidopsis thaliana NPR1 gene (AtNPR1). AtNPR1 was under the control of the constitutive CaMV 35S promoter. Natural infection by Agrobacterium to produce crown galls was observed on cannabis plants and confirmed by artificial inoculation and molecular analysis. Cannabis is naturally susceptible to Agrobacterium and in vitro transformation can lead to transgenic tissue development, providing opportunities for plant improvement through biotechnology. Further research is needed, however, to regenerate whole plants from transgenic callus.
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Thesis advisor: K., Punja, Zamir
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