Development of Egyptian barley with HMW-GS Dy10 gene of wheat

Transgenic barley plants with high-molecular-weight glutenin subunit [HMW-GS] gene from wheat were successfully generated. The HMW-GS Dy10 gene, known to be essential in bread industry, was introduced into the Egyptian barley cv. Giza 123 by biolistic bombardment. The transgenic plants, regenerated from immature embryo-derived callus cultures, were normal and fertile. Stable integration of Dy10 transgene was confirmed by molecular analysis and its expression was studied by protein analysis. Dy10 gene was co-transformed into barley with the plasmid pAHC25 harboring the bar gene for selection and the gus reporter gene. Production of Egyptian barley with wheat HMW-GS genes could be used to develop barley flour with the unique properties of wheat flour for use in bread industry

Integration and expression of the high-molecular-weight glutenin subunit Dy10 gene into Egyptian wheat

To investigate the possibility of manipulating gluten dough strength and elasticity by increasing the high molecular weight glutenin subunits [HMW-GS], bread wheat cv. Giza 164 was transformed with HMW-Dy10 subunit gene. Immature embryo-derived calli were co-transformed with a plasmid [pK-Dy10] harboring HMW-gene [Dy10] driven by its own promoter and pACH25 plasmid containing the scorable gus gene and the selectable bar gene. Integration of the three transgenes had been confirmed in the genome of transgenic T[0] plants by PCR analysis. Expression of gus gene was detected in transformed plants by histochemical staining and the expression of bar gene was detected using leaf painting assay. Grains of transgenic and non-transgenic [control] wheat plants were analyzed to estimate the level of glutenin protein using HPLC and it revealed higher levels of glutenin in transgenic grains comparing with control

simple and reliable PCR-based method for detection and screening of transgenic plants transformed with the same endogenous gene

To detect integration of a transgene in genetically modified plants that have a native copy of the gene, a simple and reliable PCR-based method has been established. This method is then used to screen for transgenic wheat plants that have been transformed with the high molecular weight glutenin subunit Dy10 gene. Two PCR primers were designed, the forward one matched upstream flanking nucleotide sequences in the pBlueScript KS vector and the reverse primer hybridizes with a Dy10 specific. Use of these primers produced a distinguishable marker only from the transgene with no appreciable signal detected from the endogenous gene