Perbandingan Tiga Metode Transformasi Agrobacterium Untuk Pencarian Gen-gen Terkait Toleransi Kekeringan Menggunakan Transposon Ac/Ds pada padi cv. Batutegi

Abstract

ABSTRACT Transformation Strategy for Indonesian Indica Rice in Attempt to Discover Drought-Tolerant Related Genes Using of Transposon Ac/Ds. Attempt to identify, isolate the gene, and study for gene function for several agronomical traits have been done including some drought tolerant traits. Japonica rice cultivars have been used due to its higher efficiencies compared with indica cultivars. Two plasmids namely pNU400 and pUR224 were used to generate mutants of these cultivars (Batutegi dan Kasalath cultivars). Those plasmids contain an element called Activator (Ac) and Dissociator (Ds) respectively. The pNU400 contains GFP (green flourescens protein) as a selectable marker, whereas the pUR224 contains hygromycine resistant gene and gusA as a reporter gene. Each plasmid was transformed into rice genome of Batutegi and Kasalath cultivars by Agrobacterium mediated transformation using three methods of transformation (A, B and C). The transformation method A was not suitable for both cultivars, where none of plantlets were produced from pNU400 and pUR224 plasmids. The transformation method B produced some plantlets from the Kasalath cultivar only using pUR224 plasmid. The transformation method C was the best method to produce transgenic plants from both cultivars (Batutegi and Kasalath), using both plasmids (pNU400 and pUR224). The PCR analysis showed that 19 and 9 plants of Batutegi and Kasalath contained both gusA and hpt genes respectively. None of those plants contained of gusA gene. Southern blot analysis revealed 3 independent lines from Batutegi dan 7 independent lines from Kasalath. The integration of Ac transposon was analyzed based on expression gfp gene when observed under UV dark reader. This research has proved that indica rice cultivars, especially the Batutegi cultivar of Indonesian origin, could be transformed. The cultivar could be used as plant model for the indica transformation. Key words: transformation, drought tolerant, indica rice, Ac/Ds transposons, Agrobacterium.