To study the in vitro invasiveness of side population (SP), cells purified from hepatoma SMMC-7721 cells and the expressions of epithelial-mesenchymal transition (EMT) markers of SP cells and non-SP (NSP) cells, fluorescence-activated cell sorting (FACS) was used to sort SP cells and NSP cells from SMMC-7721 cells. The in vitro invasiveness of both subpopulations was tested by Transwell chamber assay. Quantitative real-time reverse transcriptase-polymerase chain reaction (RT-PCR) and western blot were applied to evaluate the expressions of Vimentin, E-cadherin, N-cadherin and Snail in SP cells and NSP cells, respectively. In Transwell invasion assay, the number of SP cells (66.3 ± 7.4) penetrating the basilar membrane was significantly higher than that of NSP cells (27.5 ± 6.5) (P < 0.01). The mRNA expressions of Vimentin, N-cadherin and Snail was significantly increased in SP cells (9.527-fold, 6.834- fold and 8.173-fold, respectively) when compared with that in NSP cells (P < 0.01). In contrast, the decrease of E-cadherin mRNA expression in SP cells (5.353-fold) was markedly higher than that in NSP cells (P