| Summary: | A research report submitted to the Faculty of Science, University of the Witwatersrand,
in partial fulfilment of the requirements for the degree of Master of Science,
Johannesburg, February 1998. === Sweet potato (Ipomoea batatasy is one of the six biggest crops in the world, its high
nutritional content and large yield in tropical areas making it a useful food source,
especially in developing countries. Genetic engineering has the ability to overcome
factors such as insect and disease damage which are currently limiting its potential.
With this in mind research has been conducted into the development of a protocol to
generate transgenic sweet potato from tubers of a local South African cultivar, blesbok.
A protocol has been developed which appears capable of generating transgenic plants.
Transformation of blesbok tuber tissue was carried out by Agrobacterium tumefaciens
mediated transfer of three different binary vectors containing the uidA gene encoding
l3-glucuronidase, the npt I 1 gene conferring kanamycin resistance and the bar gene
conferring L-phosphinothricin resistance. Long term, stable expression of kanamycin
and L-phosphinothricin .resistance was confirmed with kanamycin and Lphosphinothricin
screening. Long term, stable expression of l3-glucuronidase was
confirmed with fluorescence histochemical studies employing ImaGene Red™. This
was further confirmed with quantitative assays of l3-giucuronidase activity using 4·
methylumbelliferyl-f-Dcglucuronic acid which showed an average activity of 2.82
nmole.mln'l.mg" protein. Long term, stable integration of uidA into the plant genome
was confirmed with polymerase chain reaction amplification screening. Transformed
tuber tissue was regenerated via shoot organogenesis to stem structures similarly
produced from non transformed tuber tissue. This was achieved for optimised
transformation conditions and focused on shoot induction with 2 mg.l" of the auxin
2,4-dichlorophenoxyucetic acid and 0.2 mg.l" of the cytokinin 6-benzylaminopurine.
The stems produced still need to be stimulated to develop fully into transgenic plants.
This will probably require a sharp increase in the cytokinin.auxin concentration ratio
after initial shoot induction. === MT2016
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