Evaluation of Factors Influencing In Vitro Regeneration and Transformation Protocols to Produce Salinity Tolerant Tomato (Solanum lycopersicum L.)

Ferdous, Manzur-E-Mohsina; Datta, Anamika; Islam, Aparna

doi:10.30491/jabr.2022.336979.1519

Evaluation of Factors Influencing In Vitro Regeneration and Transformation Protocols to Produce Salinity Tolerant Tomato (Solanum lycopersicum L.)

Document Type : Original Article

Authors

Manzur-E-Mohsina Ferdous ¹

Anamika Datta ²

Aparna Islam ³

¹ Department of Plant and Environmental Sciences, Copenhagen University, Copenhagen, Denmark

² Department of Stem Cell Biology, Graduate School of System Life Sciences, Faculty of Arts and Science, Kyushu University, Fukuoka, Japan

³ Department of Mathematics and Natural Sciences, BRAC University, Dhaka, Bangladesh

10.30491/jabr.2022.336979.1519

Abstract

Introduction: Tomato (Solanum lycopersicum L.) is one of the essential vegetables worldwide, for consumption and mitigating malnutrition. Genetic transformation conceivably overcomes its yield challenges due to salinity, a crucial constraint for the economical use of 30% of cultivable lands in the coastal region of Bangladesh. Therefore, a robust and reproducible protocol has been established for in vitro regeneration and transformation to develop transgenic salt-tolerant tomato plants.
Materials and Methods: During micropropagation, cotyledonary leaf explants were excised and cultured on MS media containing different combinations and concentrations of plant growth regulators. In transformation, the pre-cultured explants were inoculated and co-cultivated with Agrobacterium. Then they were transferred to the antibiotics-supplemented media to achieve salt-tolerant putative transformed plants. The transformation was confirmed by β-glucuronidase (GUS) assay and PCR for the antiporter gene.
Results: Maximum regeneration response was achieved from the explants abaxially positioned at a 1.5 cm distance apart. BARI Tomato 14 and BINA Tomato 3 showed the highest shoot regeneration response (%) on MS media supplemented with 2 mg/L BAP and with 0.1 mg/L IAA for BARI Tomato 2 and BARI Tomato 15. Bacterial culture of OD600 0.68 for 30 min and a Co-cultivation period of 48 h resulted in the highest transformation frequency (47%) in Agrobacterium-mediated transformation with pBI121 in BARI Tomato 3. The highest regeneration frequency (20.5%) was obtained in transformation with pH7WG2_OsNHX1_1.6.
Conclusions: The optimized procedure is simple, efficient, and can be used for micro-propagation and the production of tolerant varieties to increase yield in saline areas.

Keywords

Nutritional Crop

Tissue Culture

Agrobacterium-mediated Transformation

Salt Tolerance

Phytohormones

Explants Spacing