Insertion mutagenesis has become one of the most popular methods for gene functions analysis. Here we report a two-element Ac/Ds transposon system containing enhancer trap and gene trap for gene tagging in rice. The excision of Ds element was examined by PCR amplification. The excision frequency of Ds element varied from 0% to 40% among 20 F(2) populations derived from 11 different Ds parents. Southern blot analysis revealed that more than 70% of excised Ds elements reinserted into rice genome and above 70% of the reinserted Ds elements were located at different positions of the chromosome in rice. The result of histochemical GUS analysis indicated that 28% of enhancer trap and 22% of gene trap tagging plants displayed GUS activity in leaves, roots, flowers or seeds. The GUS positive lines will be useful for identifying gene function in rice.