1986 Fiscal Year Final Research Report Summary
Functional Analysis of Plasmid pSAl and Its Derivatives in Streptomyces azureus
| Project/Area Number |
60560120
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
発酵・醸造
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| Research Institution | Kyushu University |
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Principal Investigator |
OGATA Seiya Associate Professor, Faculty of Agriculture, Kyushu University, 農学部, 助教授 (20038277)
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| Co-Investigator(Kenkyū-buntansha) |
YOSHINO Sadazo Research Associate, Faculty of Agriculture, Kyushu University, 農学部, 助手 (80117291)
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| Project Period (FY) |
1985 – 1986
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| Keywords | Pock formation / Pock plasmid / Function of pock plasmid / Defective plasmid / Restriction enzyme cleavage map of plasmid / Inhibition of spore formation / Integrated state of plasmid / 胞子形成抑制機能 |
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| Research Abstract |
Streptomyces azureus ATCC 14921 (PK0) harbored one copy or less of plasmid pSA1 (8.8kb), also its primal sequences in an integrated state, and elicited pocks at 0.1 to 1% frequency on the plasmid-free strains. Strain PK100 carried 20 to 30 copies of pSA1.1 (8.8kb), a derivative of pSA1, and elicited pocks at 100% frequency. Stain PK10 carried two plasmids (pSA1.1, pSA1.2), and elicited pocks at 10 to 50% frequency. Plasmid pSA1.2 (7.6kb; copy size, 5 to 10) had two deleted portions (ca. 1.2kb and 30b long) of pSA1.1 together with no pock-forming ability. Either one or both of the deletion segments seem to be essential to the pock formation. The production of spores and thiostrepton was very low in PK100, normal in PK0 and PK10, and abundant in plasmid-free strains. These results show that the inhibition on the secondary products and the high pock formation are due to pSA1.1 in high copies. Plasmid pSA1.1 from PK10 amplified 20 to 30 copies in transformants as well as from PK100 and inhibited their spore and thiostrepton production, whereas, in PK10, copy size was lower (10 to 20) and no inhibition was observed. These results show that the defective pSA1.2 appears to depress the ability of pSA1.1 on the host cells. Cysteine also depressed the ability of pSA1.1. A specific protein (MW ca. 70,000) was found in the strains having pSA1.1.
The hybrid plasmids were easily obtained from pSA1.2 and E. coli plasmid pBR322 or pUC13 using all the unique sites of endonucleases. But, only one hybrid was obtained from pSA1.1 and pBR322 using their BamHI sites. These results indicate that the sequence of defective pSA1.2 is easily amplified in the host-vector system of E. coli but not that of pock-forming pSA1.1. These different features between pSA1.1 and pSA1.2 may be due to the 30b-deletion segments in pSA1.2 near by BamHI site.
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