Establishment Of A Multipurpose T-DNA Tagging System And Creation Of Rice Mutants Population

The main task of rice genome research has been shifted to functional genome study since rice sequencing was completed. The most direct and efficient way of functional genome research is to analyze phenotypes through a large number of mutants population generated by T-DNA tagging. In last decade, a number of T-DNA tagging system have been developed, such as gene trapping, activation tagging, multi-purpose T-DNA tagging and inducible tagging etc. Among them, multipurpose tagging is the most efficient strategy, which combines several purposes in one system.In this experiment, a collection of mutant plants of Oryza sativum cv Nipponbare was created through Agrobacterium tumefaciens-mediated transformation. The plasmid used pCAS04 was a vector with new T-DNA multipurpose tagging, in which a promoter-less GUS reporter gene was added near the right border as gene trapping and an actinl promoter including its first intron near the left border as activation tagging. A strong promoter of Maize Ubi and its first intron was used to control the selective marker nptll so as to improve the expression of the gene and to increase the efficiency of resistant calli.To achieve the best rice transformation efficiency, the transformation procedure for Nipponbare has been improved via optimization of media of each stage, co-cultivation procedure, etc.At present, more than 15 000 T-DNA tagging plants have been generated, and the GUS assay has been performed with some lines. The efficiency of gene trapping was 12.4% for roots, 7.7% for leaves, 35.5% for flowers and 10.4% for immature seeds. The patterns of GUS expression were further studied. The results obtained were important clues for functional genes research.Among the TI generation of 70 primary transgenic lines, 4 lines mutants of fertility have been found. Their phenotype was partial fertile, few fertile and sterile respectively. There also has a mutant with shape varying of the spikelet and seeds. All the mutants were show phenotype segregation, which indicate that the mutants were generated by genetic variation. The isolation of corresponding genes was in progress.