The surveillance of foodborne pathogens in food industries has shown the urgent need for rapid and dependable methods to detect and characterize the organisms in food and environments of clinical and epidemiologic importance. Recent studies on rapid methods in microbiology have been focused on biochemical characterization, immunoassays, and molecular methods. Many molecular methods have been developed and applied to the detection and characterization of foodborne pathogens in laboratories and food industries. They can be mainly divided into DNA banding pattern-based tests and DNA sequence-based tests. The former includes nucleic acid hybridization, polymerase chain reaction (PCR), amplified restriction length polymorphism, and randomly amplified polymorphic DNA, etc. Most of these methods in commercial applications are based on PCR or hybridization techniques. The principle, characteristics, and application of molecular methods for the detection and characterization of foodborne pathogens were reviewed in this article.