Browsing by Author "Klump, HH"

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    Cloning and expression of a chimeric protease inhibitor encoding gene in Escherichia coli and Pichia pastoris
    (1996) Matsebula, Aaron Mfanuzile; Botes, DP; Klump, HH; Maeder, Dennis
    Squash family protease inhibitors are small peptides of 27-32 residues, hence they are ideal subjects for structure-function studies. Their small size is within the reach of peptide chemical synthesis, which enables one to produce enough peptide material for experimental purposes within a reasonable time frame.
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    DNA based networks, nanostructures, and meta-stable states
    (2006) Lin, Chih-Ta; Klump, HH
    Nanotechnology is defined as the technology that allows to fabricate nanoscale materials and manipulate them at the molecular or atomic level. Non-biotechnology is one of the sub-disciplines of nanotechnology. Materials used for nano-biotechnology are macromolecular materials found in nature with specific mechanical and biological functions. One of the most important materials is deoxyribonucleic acid (DNA). DNA has several unique properties that make it a good candidate for nanotechnology research. The primary aim of this research is to use DNA three-way junctions as building blocks and to assemble them into a huge DNA networks through self assembly followed by DNA ligation. The second step aims of assembling six DNA three-way junctions with different sticky end sequences to from an opto-mechanical hexagonal DNA nano-switch. The switching mechanism in the righ is based on the conformational change of B- to Z- DNA. This change is influenced by the environmental cation concentrations. The third approach presented here aims to create meta-stable DNA nanostructures based on a DNA hairpin loop that can mimic the enzyme-substrate binding mechanism. This mechanism is demonstrated using a "strand displacement strategy" (SDS). Lastly, we employed a novel way of organizing hairpin loop DNA structures based on the orientation of consecutive hairpin loop DNA structures "on top" of one another.
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    TAXI : a new vehicle for the transfer of genes into monocotyledonous plants
    (1996) Chen, Wusi; Von Holt, Claus; Thomson, Jennifer Ann; Klump, HH
    The transfer of foreign genes into cereals followed by their correct expression in a tissue specific, developmentally regulated manner has become an important research focus. Based on the mechanism of Agrobacterium tumefaciens mediated transformation in dicots, a modified method to transform the monocotyledonous rye ( Secale cereale L. ) via A.tumefaciens mediated transformation was attempted. The induced bacterium culture was injected into rye seedlings, the transferred reporter gene, uid.A, was detected by PCR, and the expression of the gene was tested by histochemical assays. However, successful transformation and integration of the transgenes remained doubtful, because the frequency of kanamycin resistance in the progenies ( Rl , R2 and R3 ) did not increase. To achieve a real transformation and heritable transgenic rye, a new vehicle for gene transfer to plants was developed. A macromolecular complex, termed the TAXI, consisted of histone HI-protected single stranded DNA, containing a selectable marker gene (npt JI), linked either to a reporter gene (uidA) or a glutenin gene. The constructs were transferred by injection of rye seedlings. Molecular analyses demonstrated that all three genes were integrated and expressed in transformed rye and their progenies ( Rl and R2 ). TAXI mediated gene transfer to rye revealed an important advantage in that single or low numbers of transgenes were inserted into the transformed plant genome. However, the method of TAXI delivery to plants was not efficient. To improve this, a new approach, combining TAXI transformation and the biolistic process, was developed. A rapid regenerable callus line of a grass species, Digitaria sanguinalis, was established as a test system. TAXI coated gold particles, carrying a selectable marker gene (bar) and a reporter gene (uid.A), were used in bombardment experiments. The results of herbicide resistance and molecular analyses demonstrated that single copies or low numbers of the bar gene were inserted and expressed in regenerated transformed D. sanguinalis. Mendelian segregation in the Rl population was observed in four out of five transgenic lines.