Optimization of in vitro culture and transformation of betaine aldehyde dehydrogenase gene (BADH) into Solanum lycopersicum to produce of drought stress tolerant plant.

Document Type : Research Paper

Authors

1 Department of Plant Sciences, Faculty of Biological Sciences, Kharazmi University, Tehran, Iran.

2 Plant Bioproducts Group, Institute of Agricultural Biotechnology, National Institute of Genetic Engineering and Biotechnology, Tehran,

3 Department of Plant Sciences, Faculty of Biological Sciences, Kharazmi University, Tehran, Iran

4 Central Branch of Islamic Azad University, Tehran, Iran

Abstract

Tomato (Solanum lycopersicum L.), a major Solanaceae crop, is sensitive to drought and osmotic stress. Enhancing tolerance via genetic transformation is promising for sustainable production. This study examined two cultivars, CH and Rio Grande, optimizing in vitro regeneration and introducing the spinach betaine aldehyde dehydrogenase (BADH) gene, which aids glycine betaine biosynthesis, an osmoprotectant. Cotyledonary and hypocotyl explants from 10-day-old seedlings were cultured with varying benzyl adenine (BA) and indole acetic acid (IAA) concentrations. In CH, cotyledons showed 96.26% regeneration and hypocotyls 92.53% on 3 mg l-1 BA with 0.1 mg l-1 IAA. Rio Grande had 73.0% regeneration in cotyledons at 2.5 mg l-1 BA + 0.1 mg l-1 IAA and 66.6% in hypocotyls at 3 mg l-1 BA. Cotyledons produced more shoots consistently. Shoot multiplication improved with gibberellic acid (GA3) + IAA; 3 mg l-1 GA3 and 0.1 mg l-1 IAA yielded 6.67 and 6.33 shoots per cotyledon explant in CH and Rio Grande, respectively. Agrobacterium tumefaciens carrying pRI201-AN with BADH infected cotyledons for 30 seconds, followed by 48-hour dark co-cultivation at 28°C with 40 mg l-1 acetosyringone, maximizing transformation. PCR confirmed a 1668 bp BADH transgene band in resistant plants. Under mannitol-induced osmotic stress at –6 bar, controls showed chlorosis and death, while transgenic plants survived and grew, demonstrating enhanced osmotic tolerance. This study shows optimized regeneration and BADH gene transfer improved tomato osmotic stress tolerance, supporting BADH's role in breeding stress-resilient tomatoes.

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