Genetic transformation of Streptomyces caespitosus

Genetic transformation of Streptomyces caespitosus by plasmid pIJ 702 was carried out. Optimal conditions for the protoplast preparation of Streptomyces caespitosus, its regeneration, and its transformation by pIJ 702 were evaluated. Addition of 2% glycine to the culture broth was optimal for protoplast yield. Formation and regeneration of protoplasts were most efficient when the mycelium were harvested at between late log and stationary growth phase. The regeneration frequency of the protoplasts was 15% when the protoplasts were regenerated on R2YE agar media containing 0.5M sucrose. Under the best condition for protoplate-regeneration, the optimal transformation frequency was achieved with 40% polyethylene glycol (M.W. 4,000) treatment for 2 minutes.ope