Agrobacterium-Mediated Transformation of Indica Rice: A Non-Tissue Culture Approach

Gramineous crop plants including important cereals like rice (Oryza sativa L.) were classified as recalcitrant to transformation because of their both low morphogenic potential and transformation efficiency. An efficient in planta transformation method, via physical wounding of the embryo region was conducted in two rice landraces (O. sativa ssp. Indica) where the seeds were soaked in water for 48 hours and pierced with an Agrobacteriumcoated needle. Two strains of A. tumefaciens (EHA105 and LBA4404) harbouring pCAMBIA1105.1r vector were used. A three step method in cell culture was applied for vir gene induction in Agrobacterium. Integration and presence of the transgene into the genome of putative transgenic rice plants was confirmed using polymerase chain reaction, resistance of leaf tissues to hygromycin and histiochemical GUS assay. In a comparative study, EHA105 strain in the presence of 100 ?M acetosyringone on ‘Hashemi’ cultivar had a significantly higher transformation efficiency (~9.0%) compared to the other treatments. Ninety T0 seedlings were selected randomly and analysed further at T1 generation, which ~28% confirmed to be positive.