Genetic Transformation of Immature Zygotic Embryos of Maize Genotypes via Agrobacterium tumefaciens
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Many cereals crops are recalcitrant species to genetically modification through their resistance to Agrobacterium infection and recalcitrance to in vitro regeneration. However, a routine and efficient transformation protocol of Syrian maize (Zea mays) using an Agrobacterium tumefaciens standard binary vector system for year round production of fertile transgenic maize plants was achieved. Immature zygotic embryos of Syrian genotypes and the control hybrid line Hi II were infected with A. tumefaciens strain EHA101 harboring a standard binary vector pTF102. The average stable transformation frequency (number of bialaphos-resistant events recovered per 100 embryos infected) of the present protocol was 14.5% for Hi II and 5.9 for Syrian genotypes. The expression of the beta-glucuronidase (GUS) gene was delivered with high efficiency to maize calli, roots and shoots by A. tumefaciens carrying the GUS gene was observed. Progeny analysis through polymerase chain reaction (PCR) analysis of total isolated DNA confirmed the integration, expression and inheritance of T-DNA carrying the selectable marker gene bar and reporter gene gus in the genomes of transgenic maize plants. More than 90% of transformants were normal in morphology. The protocol took about 3 months from the start of infection and cocultivation to the planting of transformants into pots. It is anticipated that this study will assist further enhancement of maize transformation technology leading to develop an updated protocol for the Agrobacterium-mediated generation of maize. To our knowledge, this is the first report providing evidence of the transformation ability of Syrian genotypes via A. tumefaciens. [Ayman A, Lane S, Fuller MP.
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