Three gene clusters controlling the biosynthesis of the antibiotics actinorhodin (ACT),undecylprodigiosin (RED) and calcium-dependent antibiotic (CDA) were sequentially knockouted by gene replacements from the chromosome of Streptomyces lividans TK24 to derive SBT5.In SBT5,the global regulatory genes afsRScla from Streptomyces clavuligerus was integrated in the place of the cda gene cluster.The act gene cluster from Streptomyces coelicolor was introduced into SBT5 to test its capability of expressing secondary gene cluster.The resulting SBT5 exconjugants overproduced ACT comparing with wild-type strains harboring double copies of the act gene cluster.SBT5 is an efficient host with a clean secondary metabolism background suitable for heterologously expressing and screening secondary metabolic gene clusters.