Balakrishnan, Bhavani
(2013)
Preliminary Assessment Of Conditions Influencing Agrobacterium Rhizogenes- Mediated Transformation Of Eurycoma Longifolia Jack. Somatic Embryos.
Masters thesis, Universiti Sains Malaysia.
Abstract
Effective propagation of in vitro callus, shoot, plantlet and somatic embryo cultures of Eurycoma longifolia was achieved using micropropagation technique. In vitro shoots, somatic embryos and plantlet cultures were used for conducting histological and SEM analysis, chemotaxis assays and transformation with four different Agrobacterium rhizogenes disarmed strains. Histological analysis and SEM observations revealed that somatic embryos of Eurycoma longifolia share similar traits with zygotic embryos. Positive chemotactic responses of A. rhizogenes towards somatic embryo explants were detected using swarm plate assay. In vitro shoot and plantlet cultures were used to obtain leaves, stems and rachis explants for transformation. Hairy root-like structures were seen emerging from in vitro stem explants co-cultivated with Agrobacterium rhizogenes strain AR14. However, the number of explants expressing these hairy root-like structures was too low to be confirmed using GUS assay or PCR based detection of transgenes. Transformation optimisations with somatic embryo explants were conducted by analysing the effect of four different factors on the GUS positive frequency. It was ascertained that AR12 is the most virulent and suitable Agrobacterium rhizogenes strain for transformation of E. longifolia somatic embryos.
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