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Sorghum (Sorghum bicolor L. Moench) ranks fifth, worldwide in production among cereals and it is unique in that
it is adapted to semi-arid environments. Sorghum grain with enhanced protein quality could contribute significantly
to improving the nutritional value of the diets of people and livestock dependent on sorghum as a major protein
source. The proposed research aims at genetically enhancing the nutritional quality of grain sorghum by the
introduction of genes encoding the methionine-rich maize beta-zein and the lysine-rich barley chymotrypsin inhibitor
CI-2 proteins, with the goal to produce transgenic sorghum plants with elevated lysine and methionine contents.
Development of an efficient routine transformation protocol for grain sorghum
An efficient transformation protocol for sorghum would form the technological basis for nutritional quality
improvement of grain sorghum. As a first step towards achieving this goal, two transformable sorghum genotypes
were selected, and five African sorghum genotypes are currently being assessed on various tissue culture media
compositions to identify additional highly regenerable and transformable genotypes. Particle bombardment experiments
optimising parameters for stable transformation of sorghum using either the commercial BioRad gun or the Particle
Inflow Gun were done. Furthermore, two vectors were prepared for Agrobacterium-mediated transformation of grain sorghum.
Methionine and lysine rich protein genes
The ultimate goal of this project is the production of transgenic grain sorghum with improved nutritional value
due to elevated lysine and methionine contents. Four constructs containing genes encoding either methione-rich or
lysine rich proteins, driven by strong seed specific promoters, were prepared for particle bombardment-mediated
transformation of grain sorghum. Two constructs containing the wild type CI-2 gene driven by the maize gamma-zein
(?-zein) promoter were prepared with signal peptides with one also containing a c-terminal KDEL ER-retention signal
to increase protein stability. Two constructs were prepared containing the methionine-rich beta-zein gene or fusion
protein gene driven by the HMW or gamma-zein promoter, respectively. In order to detect the recombinant protein in
the future transgenic sorghum, an epitope tag was fused in frame to the methionine-rich protein genes. In addition,
engineering of the lysine-rich wild type CI-2 gene, with additional lysine substitutions in a reactive loop region,
is underway.
The plant expression vectors containing the reporter gene uidA (GUS) and selectable marker gene hpt II, as well
as the lysine-rich CI-2 gene under control of the gamma-zein promoter were prepared for Agrobacterium-mediated
transformation of selected sorghum genotypes.
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