Protocol

Transgenic Plants: Methods and Protocols

Volume 286 of the series Methods in Molecular Biology™ pp 141-150

Organogenesis From Transformed Tomato Explants

  • Anne FraryAffiliated withDepartment of Molecular Biology and Genetics, Izmir Institute of Technology
  • , Joyce Van EckAffiliated withThe Boyce Thompson Institute for Plant Research

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Summary

Tomato was one of the first crops for which a genetic transformation system was reported involving regeneration by organogenesis from Agrobacterium-transformed explants. Since the initial reports, various factors have been studied that affect the efficiency of tomato transformation and the technique has been useful for the isolation and identification of many genes involved in plant disease resistance, morphology and development. In this method, cotyledon explants from in vitro-grown seedlings are precultured overnight on a tobacco suspension feeder layer. The explants are then inoculated with Agrobacterium and returned to the feeder layer for a 2-d period of cocultivation. After cocultivation, the explants are transferred to an MS-based selective regeneration medium containing zeatin. Regenerated shoots are then rooted on a separate selective medium. This protocol has been used with several tomato cultivars and routinely yields transformation efficiencies of 10–15%.

Key Words

Agrobacterium tumefaciens biotechnology gene transfer genetic engineering genetic transformation Lycopersicon esculentum morphogenesis regeneration transgenic plants