The biological activity of polyketide antibiotics is often strongly dependent on the presence and type of deoxy sugar residues attached to the aglycon core. In this study, an efficient host-vector system was developed based on the pikromycin producing strain of Streptomyces venezuelae for combinatorial biosynthesis of deoxysugar moieties and its attachment to the aglycon substrate. First, the S. venezulae mutant strain YJ01 was created by deletion of the des gene cluster, desI-VII and desR, which is responsible for the biosynthesis of desosamine. DesIII and IV genes which are involved in biosynthesis pathway of various deoxysugars were integrated into the genomic DNA using an integrative plasmid. The glycosyltransferase desVII gene and genes involved in the biosynthesis of dTDP-_D-olivose were cloned into a high-copy plasmid for expression in S. venezuelae.