Rapid production of transgenic sugarcane with the introduction of simple loci following biolistic transfer of a minimal expression cassette and direct embryogenesis

A protocol is described that supports the production of transgenic sugarcane plants ready for transfer to soil within 3 mo from culture initiation. Biolistic gene transfer into cross-sections of immature leaf whorl explants followed by direct somatic embryogenesis resulted in the stable genetic transformation of the commercially important sugarcane cultivar CP 88-1762. Accelerating the production of transgenic sugarcane plants not only saves time and effort but will likely also minimize somaclonal variation. Southern blot analysis revealed simple transgene integration patterns ranging from one to five hybridization products. NPTII-ELISA confirmed that most of the transgenic plants expressed the transgene stably in vegetative progeny. Using a minimal, linear expression cassette (MC) without vector backbone sequences for the biolistic gene transfer and reducing the amount of MC to 10 ng per shot may have led to simple transgene integration and stable transgene expression. Therefore, this protocol has great potential for the generation of commercial transgenic sugarcane events.