Promoter Trapping And T-DNA Tagging In Rice (Oryza Satova L.)

Rice (Oryza sativa) is one of the most important grain cropsin the world. It is also a model for molecular study of cereals.With the completion of rice genome sequencing, rice genomeresearch has been shifted to functional genome study. One of thestrategies for functional genome study is to use a largepopulation of mutants generated by T-DNA tagging.In this work, T-DNA insertional mutants were produced throughAgrobacterium-mediated transformation and make some analysis ofimportant mutants. The pAHCL used in this work is a promotertrapping vector which has apromoterless GUS reporter gene locatednear the right border. In total, 393 T-DNA tagging plants wasgenerated. PCR and Southern blot analysis showed that thehygromycin resistant lines were transformants, and the copynumber of the T-DNA ranged from 1 to 4 in different lines. TheGUS assay on 393 lines showed the efficiency of gene trapping was5.34% for leaves, 1.78% for roots, 10.17% for endosperms, 6.36% forcoleoptiles, respectively. 8 T-DNA flanking sequences from 12 GUSpositive lines were obtained by PCRWalking.Further analysis of the T1 generation of 65 T-DNA taggingplants was performed. 18 of 65 T-DNA tagging lines showed markablemutant characters: dwarf, semi-dwarf, sterile, half sterile,glume blaze, single tiller, multitiller, small spike, rolled leaf.The T-DNA flanking sequences were obtained from 5 mutants by Genome Walking. The chromosome location of flanking sequences andpossible encoding proteins were presumed by homologous search inthe rice database in NCBI.