Transgenic Plants of Markerless Antisense Waxy Indica Rice Obtained by Agrobacterium-Mediated Co-transformation

【Objective】The antisense Waxy gene was transformed into the mature indica rice embryo calli by the transgenic technology in order to improve the amylose content in indica rice. 【Method】The plasmid p13W8 carrying antisense fragment of Waxy gene and plasmid pCAMBIA1300 containing hpt gene were introduced into the calli derived from mature indica rice embryo by Agrobacterium tumefaciens-mediated co-transformation. 【Result】The results showed that the calli from the seed embryo and subcultured for 18-24 d had higher transformation and differentiation frequency and could provide the acceptor for the Agrobacterium-mediated co-transformation. hpt-positive frequency was 97.6% and Waxy-positive frequency was 27.1% in the hygromycin-resistant calli by PCR analysis. Five plants showed positive bands for Waxy gene by PCR analysis in 46 transgenic hpt-positive plants, the Waxy-positive plants frequency was 10.9%. The segregation of antisense fragment of Waxy gene and hpt gene was observed by PCR analysis in four T1 population. Thirty-nine plants containing only the antisense fragment of Waxy gene were identified in 286 T1 plants, the Waxy-positive plant frequency was 13.6%. The amylose content of some seeds derived from transgenic plants with only the antisense fragment of Waxy gene was found with different varying degree of reduction. 【Conclusion】The indica rice plants only with antisense Waxy gene and amylose content decreasing significantly were obtained by Agrobacterium tumefaciens- mediated co-transformation.