Transformation and expression of a gene for herbicide resistance in a Brazilian sugarcane

Embryogenic calli of the Brazilian sugarcane (Saccharum officinarum L.) gen otype SP80-180 were transformed with two plasmids containing genes coding f or neomycin phosphotransferase (neo) and phosphinotrycin acetyltransferase (bar), by particle bombardment using an apparatus developed at Copersucar T echnology Center. Transformed plants were initially selected on culture med ium containing Geneticin, and resistance was confirmed by localized applica tion of a kanamycin solution to leaves of hardened plants at the nursery. A commercial formulation of ammonium gluphosinate was sprayed twice on these antibiotic-resistant plants. The resistant plants were considered co-trans formed, and Southern analysis confirmed stable integration of both bar and neo genes. In addition, phosphinotrycin acetyltransferase expression was su pported by RT-PCR analysis and neomycin phosphotransferase presence was dem onstrated by western blotting. Similar analyses were also performed with mi cropropagated transformants after three cycles of subculture.