IMPROVEMENT OF TRANSFORMATION SYSTEM IN STREPTOMYCES USING A MODIFIEDREGENERATION MEDIUM

For improvement of the regeneration frequency of Streptomyces griseus 2247 protoplasts, the effects of growth phase and components of the re generation medium on the frequency were investigated. Protoplasts prep ared from transition-phase mycelia of S. griseus 2247 exhibited the hi ghest regeneration frequency. An improved regeneration medium designat ed R1M was established by supplementing Polypepton and yeast extract w hich were shown to increase the S. griseus 2247 growth rate to R1 prev iously developed for S. griseus. The regeneration frequencies of S. gr iseus 2247 and its pleiotropic mutant HT-3 were greatly increased to 8 7.5% and 92.5%, respectively, on R1M medium. Using this protoplast-reg eneration system, the transformation efficiency for the plasmid pRES18 was increased to 10(6) transformants per mu g DNA. R1M medium was als o effective for the protoplast regeneration of some other antibiotic-p roducing streptomycete species.