The cryptic low copy plasmid designated as plMB R8 was isolated from an industrial strain of the chlortetracycline producer Strep/onr~ces ozireo/Ucirns R8/26. Using restriction endonucleases the pIMB R8 plasmid was characterized to have 15 kb. Subcloning of the Bar17 HI fragments of plMB R8 into rcplication probe vector resulted in the identification of the rcplication part. The 0.7 kb Be11 -Byfl l'ragnicnt is sufficient for normal replication. but produces about fourty times high plasmid copy numbers than the original pIMB R8. Using the replication part of pIMB R8 we constructed several shuttle cloning vectors for the E. di-strcptomycete system.
Strc~ptoniyccls uureoj~ciozs is the producer of medically and commercially important antibiotics, tetracycline and chlortetracycline.
To study the biosynthetic pathway in our strain we decided to construct an E. coli-streptomycete shuttle vector system based on the pIMB R8 plasmid replicon, since some procedures can be performed much more easier in E. coli as in Sireptotnycctes. Advantages of procedures available in E. coli have been reported by LAKSON and HERSHBERCER (1984,1986), STANC'ZAK el nl. (1986) and PARADISO et d. (1987).
The essential DNA region for stable replication is often relatively small (SHINDOH i '