Abstract: Denaturing gradient gel electrophoresis (DGGE) analysis was recently applied in the detection of food microbial diversities as well as of environmental microbial communities widely. To analyze the component and structure of lactic acid bacteria in food samples and dairy fermented products, DGGE conditions such as gradient of denaturant and run time for electrophoresis were firstly optimized. The results showed that 30%～55% was a suitable denaturant range of gradient for partially melting DNA fragments, and about 200 min was a proper time for the variable V3 region separation of 16S rRNA genes of lactic acid bacteria. Five chromosomes DNA mixture of lactic acid bacteria was used as template DNA for traditional PCR and touch down PCR to amplify the V3 region, respectively. The PCR production was subjected to DGGE analysis as described above. The different efficiencies of amplification during PCR by universal bacteria primer set were also observed. This suggested that the limit in the detection potential of PCR-DGGE is a consequence of DNA template competition during the amplification reaction.

Key words: denaturing gradient gel electrophoresis, polymerase chain reaction, lactic acid bacteria, variable V3 region