Expression of synthetic Alanine Aminotransferase gene improves nitrogen uptake and response in transgenic tobacco

Abstract

Improvement in nitrogen use efficiency of crop plants has been a major challenge in agricultural research. A barley alanine aminotransferase (HvAlaAT) gene was codon optimized and got synthesized to achieve optimal expression in tobacco plants. Synthetic alanine aminotransferase (co-AlaAT) gene was cloned under 2XCaMV35S promoter and transformed in tobacco through Agrobacterium-mediated method. Out of seven independent transgenic tobacco events, three putative transgenic tobacco lines were selected for evaluation in pots at 0, 75, 100 and 125 kg N/ha nitrogen fertilizer doses applied in three split applications. Molecular analysis of the putative transgenic tobacco plants was conducted through PCR and real-time quantitative PCR. Transgenic tobacco lines evaluated through biochemical and morpho-physiological means showed improvement in N uptake, total free amino acid, crude proteins, AlaAT activity, leaf area, stalk diameter, above the ground fresh mass, above ground dry matter and seed weight per plant. All these attributes demonstrated improved nitrogen uptake and its incorporation into biomass as compared to the wild-type. From these findings, it is concluded that co-AlaAT gene could be a possible candidate for developing nitrogen responsive and N use efficient crop plants to reduce nitrate contamination in the environment and to minimize the farmer's input in terms of nitrogenous fertilizer application.