Many invasive bacteria produce capsular polysaccharides, which are essential virulence factors for pathogen invasion to human body. Vaccination using capsular polysaccharides conjugated to a protein carrier is a powerful approach for protecting humankind against infectious diseases caused by bacterial pathogens, such as Streptococcus pneumoniae and Haemophilus influenzae type b (Hib). Sufficient amounts of pure and well-defined polysaccharides, which are not easily obtained by purification or purely organic synthesis, are important to reveal their biological roles. Therefore, producing structure-defined synthetic capsular polysaccharides by following their biosynthetic pathway is an alternative and efficient solution. Furthermore, unravel of the biosynthetic pathway of the bacterial polysaccharides will open a door for further modification and improvement of polysaccharide vaccines by combinatorial biosynthesis (producing novel polysaccharides by switching genes in the capsular polysaccharide loci amongst populations of same/related species), unnatural substrate substitution, and/or chemical modification. The relationship of the capsular polysaccharide with the host immunogenicity can be further exploited using the produced novel synthetic polysaccharide products.