Microbial reduction of perchlorate in the presence of zero-valent iron (ZVI) was examined in both batch and column reactors to assess the potential role of iron as the electron donor for biological perchlorate reduction process. Anaerobic microbial mixed cultures completely removed aqueous perchlorate in the presence of zero-valent iron. In order to identify the population responsible for perchlorate reduction in the ironsupported culture, community analyses comprising of microbial fatty acids and PCRDGGE techniques were performed for perchlorate reducing cultures. Analysis of fatty acid methyl esters (FAMEs) and subsequent principal component analysis (PCA) showed a clear distinction between iron-supported perchlorate reducing culture and perchlorate-reducing cultures receiving different electron donors. Nested PCR was performed to amplify chlorite dismutase (cld) gene that is unique to perchlorate reducing bacteria (PRB) and the denaturing gradient gel electrophoresis (DGGE) was conducted to examine the effects of different electron donors on the DGGE patterns. The multiple-aligned sequences generated from DGGE fragments were compared using a phylogenetic tree. Results obtained from PCR-DGGE techniques along with phylogenetic sequence analysis elucidated (1) dominant organisms and mechanisms in the iron-supported perchlorate reducing culture, (2) genetic affiliation among PRB fed with different electron donors, and (3) the role of functional genes on microbial population dynamics. These results lead us to further understand the function of zero-valent iron as an electron source for microbial perchlorate reduction.