Abstract: DNA fragment introduced into cells can specifically inhibit the expression of homologous gene, which is identified as DNA interference (DNAi). DNAi was discovered with the events of post-transcriptional gene silencing in tobacco, and subsequently explored in other plants and animals. DNAi was present in prokaryotes and in vitro DNAi was also achieved by Ago proteins from prokaryotes. The mechanism of DNAi in prokaryotes is mostly that Ago proteins are complexed with small DNA guide to cleave DNA or RNA. Post-transcriptional gene silencing, genome methylation and transcriptional repression of promoter binding might be also covered in the mechanisms of DNAi in eukaryotes. In addition, the further research and application of DNAi were discussed and prospected.