Abstract： Objective To perform molecular evolution analysis of mazEF gene in genome sequenced strains of Escherichia coli and Shigella. Methods Pathway Tools (version 13.5) provided by BioCyc was used to search encoding gene mazF of toxin MazF ( chpA) and encoding gene mazE of antitoxin MazE (chpR) in genome sequenced 10 strains of Escherichia coli, 6 strains of Shigella and 1 strain of unkown Enterobacteria. Then Minimum Evolution method in MEGA4. 1 software was used to analyze molecular evolution of MazE and MazF. Results Encoding gene mazF of toxin MazF was found in 12 strains, and encoding gene mazE of antitoxin MazE was found in 11 strains, while neither mazE nor mazF was found in rest 5 strains. Both mazE and mazF had good conservation in molecular evolution analysis. Conclusions MazEF is the first toxin-antitoxin system found in prokaryotic chromosomes, but not in all strains of Escherichia coli and Shigella. MazEF deletion is associated with antibiotic resistance and it also mediates programmed cell death in bacteria, so MazEF might be a new target for antimicrobial agents.