Maintaining current, relevant curriculum in undergraduate Food Microbiology courses is essential for training future experts in food quality and safety. Having an understanding of the fundamental techniques (for example, polymerase chain reaction [PCR]) that are used in the food industry and regulatory agencies is critical for students entering the workforce. The purpose of this study was to assess the efficacy of integrating molecular methods into an undergraduate Food Microbiology course in both lecture and laboratory settings. Modules on PCR and pulsed-field gel electrophoresis (PFGE), both of which are currently used by government agencies and the food industry to investigate the presence and persistence of foodborne pathogens, were developed, introduced, and evaluated among 269 students over 4 y. Multiple teaching and learning styles were incorporated through (i) traditional lecture format on the basics of PCR and PFGE; (ii) hands-on group activities to build upon the lecture instruction; (iii) performing PCR and PFGE in the laboratory; and (iv) group discussions to analyze results from laboratory exercises. Pre- and postinstruction evaluations revealed significant increases in understanding and application of both methods in lecture and laboratory settings as demonstrated by 0.60 and 0.51 mean normalized gains for respective PCR and PFGE lectures and 0.50 and 0.56 mean normalized gains in respective labs. Academic year significantly impacted score improvement, potentially due to hidden factors, such as previous exposure to material and student aptitude. This study provides the platform for successful introduction of molecular techniques in an undergraduate Food Microbiology course. The guidelines and materials developed by our group are publicly available for use by other institutions.