O-antigen (O-polysaccharide) is a part of the lipopolysaccharide component of the outer membrane of Salmonella and consists of oligosaccharide repeats (O units). It is one of the most variable cell constituents and the diversity is a common basis for bacterial serotyping. Genes, which are responsible for O-polysaccharide synthesis and translocation, exist in Salmonella genome into cluster, called O-antigen gene cluster. The presence of O antigen plays a role in bacterial virulence and is essential for invasion, survival and colonization of bacteria in their natural environment. Besides, O-antigen is also a major protective antigen of Salmonella, and it can stimulate the host to produce strong immune responses, which contribute to bacteria elimination. Thus, it has extensively been targeted for vaccine development. This review summarizes O-antigen in terms of gene structures, synthesis, function and its application in vaccine development.