The repetitive element sequence-based polymerase chain reaction or rep-PCR is an effective DNA fingerprinting technique to generate the DNA profiles or fingerprints for individual microbial strains by using specific primers designed to target the repeated palindromic sequences of bacterial DNA. The amplified products are further separated by electrophoresis technique for detection of different microbial strains. The genomic fingerprint can further be employed to distinguish the sub-species level of microbial world. The application of rep-PCR has exhibited itself as a potential epidemiological tool in investigating foodborne intoxications, differentiating microbial subtypes, and analyzing microbial ecology. This protocol mainly emphasizes on the detection of Gram-positive bacterial toxins by rep-PCR in foodborne illnesses.