Efficiency of Callogenesis and Amenability to Agrobacterium Mediated Transformation of Selected Traditional Sri Lankan Rice Varieties

Abstract

Traditional rice (Oryza sativa L.) varieties possess many important traits such as medicinal and nutritional properties, drought and pest resistance, salt tolerance, etc. Efficient plant regeneration culture and transformation protocols are important in utilizing such genes. Therefore, the aim of this study was to optimize an efficient protocol for callus induction and transformation of selected Sri Lankan traditional rice varieties. Dahanala, Kaluheenaty, Kalubalawee, Madael, Kahatawalu, Kaharamana, Gonabaru, Alwee, Wannidahanala, Kottiyaran, BG380 and H4 varieties were selected for the study. Twelve different concentrations of 2,4-D and kinetin combinations in MS basal medium were used to determine the effect of above hormone concentrations on callus induction. Calli were transformed using Agrobacterium mediated transformation. For calli transformation Agrobacterium tumefaciens GV3101 containing pB1121 vector harboring GUS gene was used. Efficiency of transformation through GUS expression was calculated for each variety. There was a significant difference in response with respect to 12 treatments used for callus induction of tested varieties except Gonabaru. Varieties Kaluheenaty, Madael, Gonabarur Kottiyaran and H4 have shown the best callus induction for the same combination of hormones (2.5mg/L 2,4-D and 0.5 mg/L kinetin). Other six varieties showed best callus induction for different combinations with majority of them being in the range of 2.5mg/L 2,4-D and 0.5 mg/L kinetin. In accordance with previous work on indica rice regeneration, our results also showed that callus induction was genotype dependant. Higher GUS expression efficiencies starting from 100% were reported for most varieties. Results of this study indicate the possibility of using optimized hormone concentrations and Agrobacterium mediated transformation for selected Sri Lankan traditional rice varieties in future genomic studies.