Post-transcriptional gene silencing (PTGS) has been shown to occur in many transgenic dicotyledonous plants. Although transgene silencing has been documented in transgenic monocots, PTGS has been only recently described in monocotyledonous plants. To study PTGS in rice, we generated 25 transgenic rice cell lines in which the 35S-gus gene was stably integrated. In one transgenic cell line, lack of gus expression was shown to be caused by PTGS based on the results of run-on transcription assays. Furthermore, to examine whether reintroduction of the same plasmid DNA into cells in which the gus gene was highly expressed causes the suppression of the gus expression, the gus plasmid was introduced into protoplasts isolated from a high gus-expressing line. Results of the kinetic analysis of the GUS activities in transfected protoplasts indicated that the gus expression in transfected protoplasts was suppressed by the gus plasmid but not by the luc plasmid harboring the 35S-luciferase gene, suggesting that suppression of the gus expression by introduced plasmid DNA was homology-dependent. These results suggest that the transgenic protoplasts may be a useful experimental system to study PTGS in rice and other species.