Establishment of Agrobacterium-Mediated Genetic Transformation in Popular Rice Cultivar BRRI Dhan-29 through Reporter GUS Gene Expression System in Bangladesh

Abstract

Agrobacterium-mediated genetic transformation was done in BRRI dhan-29 using Agrobacterium strain LBA4404 harboring the plasmid pBI121. The plasmid carries CaMV35S promoter driving the expression of GUS and Kanamycin resistant genes. The β-glucuronidase (GUS) gene served as a reporter gene in the histochemical assay and the neomycin phosphotransferase II (nptII) gene used for the identification of resistance to kanamycin. The incubation period of 30 minutes was found to be the most effective for infection and integration of foreign genes into callus. Transformed calli were selected along with a control in the regeneration medium containing 100 mg/l of kanamycin. Embryogenic calli were found to show the best transformation ability producing 30 % GUS positive result in histochemical assay after 3 days of co-cultivation. GUS expression was also observed in the leaf and root of putative plants prior to final confirmation of transformation through PCR amplification.