Factors influencing Agrobacterium-mediated genetic transformation efficiency in wheat.

Abstract

Assessment of the efficiency of Agrobacterium tumefaciens mediated transformation in four bread wheat varieties was focused and upon optimization of experiments done to obtain tangible results. The four varieties namely Fielder, Navojoa, Baj and Kachu/saul were transformed with optimized protocols, subsequently randomly selected from selection medium amalgamated with phosphinothricin (Glufosinate). Initially the variety Fielder was proceeded with 36 different concentrations of synthetic auxins 2,4-D and Picloram as 36 treatments. The treatment 30 (48 g of 2,4-D and 64 g of Picloram) has shown higher efficiency and is comparable in terms of the three traits studied. Additionally, antioxidants and growth regulators were adjusted to obtain better result towards reasonable transformation frequency. Based on the optimization for the variety Fielder, additional varieties were also tested and selected against the selectable marker PPT. Selected genotypes were subjected for gusA or Bar gene PCR amplification. The result revealed the frequency of transgenesis with the range of 60-70% with the varieties studied, however the whole experiment needs further repetitions for confirmatory result. Transgenic genotypes of the Fielder variety showed moderate to strong GUS expression in leaves, anthers, seeds, seed coats, roots compare to non-transgenic control plant tissues. PPT leaf painting assay showed the lack of necrosis on painted area of leaves in transgenic genotypes compared to the non-transgenic ones.