Abstract
BackgroundThe main aim of this study was to improve fungal resistance in bread wheat via transgenesis. Transgenic wheat plants harboring barley chitinase (chi26) gene, driven by maize ubi promoter, were obtained using biolistic bombardment, whereas the herbicide resistance gene, bar, driven by the CaMV 35S promoter was used as a selectable marker.ResultsMolecular analysis confirmed the integration, copy number, and the level of expression of the chi26 gene in four independent transgenic events. Chitinase enzyme activity was detected using a standard enzymatic assay. The expression levels of chi26 gene in the different transgenic lines, compared to their respective controls, were determined using qRT-PCR. The transgene was silenced in some transgenic families across generations. Gene silencing in the present study seemed to be random and irreversible. The homozygous transgenic plants of T4, T5, T6, T8, and T9 generations were tested in the field for five growing seasons to evaluate their resistance against rusts and powdery mildew. The results indicated high chitinase activity at T0 and high transgene expression levels in few transgenic families. This resulted in high resistance against wheat rusts and powdery mildew under field conditions. It was indicated by proximate and chemical analyses that one of the transgenic families and the non-transgenic line were substantially equivalent.ConclusionTransgenic wheat with barley chi26 was found to be resistant even after five generations under artificial fungal infection conditions. One transgenic line was proved to be substantially equivalent as compared to the non-transgenic control.
Highlights
The main aim of this study was to improve fungal resistance in bread wheat via transgenesis
Polymerase chain reaction (PCR) Genomic DNA was extracted from putative transgenic plants, which were resistant to the herbicide BastaTM, and the non-transgenic
Wheat transformation and recovery of transgenic families The plant expression vector pBarley/chi/bar (Fig. 1) containing the barley chi26 gene was transfected to the immature embryos of wheat cultivar Hi-Line by particle bombarding
Summary
The main aim of this study was to improve fungal resistance in bread wheat via transgenesis. Resistant wheat cultivars can be obtained via genetic transformation, and deployed to reduce yield losses and protect seed quality deteriorated by rust and powdery mildew diseases. The expression of chitinase gene has been found to enhance the resistance against fungal diseases in many plant species via genetic transformation, for instance, the expression of a class II chitinase in Brassica juncea could successfully provide protection against Alternaria leaf spot disease [13]. Earlier studies in wheat indicated that the constitutive expression of class II barley chitinase could enhance resistance against Erysiphe graminis [17] and Fusarium graminearum [18, 19]. The aim of the present work was to evaluate transgenic wheat lines, harboring the barley chi gene for resistance against rust and powdery mildew diseases. The most promising transgenic family was analyzed in contrast to the non-transgenic controls to substantiate the resistance
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