A highly efficient method for Agrobacterium mediated transformation in elite rice varieties (Oryza sativa L. spp. indica)

Abstract

An Agrobacterium mediated transformation method was developed for the Thai rice variety,  Pathumthani 1 (PT1), and the Indian rice variety, Pokkali (PKL). Various aspects of the transformation method, including callus induction, callus age, Agrobacterium concentration and co-cultivation period were examined, in order to improve transformation efficiency. Optimized transformation conditions were established using Agrobacterium strain EHA105, which carries a virulent plasmid, pCAMBIA1301.  A modified Murashige and Skoog (MS) medium supplemented with 1 mg/l 2, 4-D and 0.5 mg/l picloram was optimized for callus induction. Three week old calli were used to co-cultivate with 0.8 -1 OD₆₀₀  Agrobacterium for 30 min and the culture was continued on agar medium without antibiotics for 2 days. This method can be used to induce high quality calli within three weeks. Based on GUS determination, it was demonstrated that the transformation method was improved significantly, with a high level of transformation efficiency.

Keywords: Agrobacterium tumefaciens, indica rice, mature seed-derived callus, rice transformation, transgenic rice

African Journal of Biotechnology Vol. 9(34), pp. 5488-5495, 23 August, 2010