1997 Fiscal Year Final Research Report Summary
Conjugal transfer of oncogenes from bacteria to animal celIs
| Project/Area Number |
08044292
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| Research Category |
Grant-in-Aid for international Scientific Research
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| Allocation Type | Single-year Grants |
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| Section | Joint Research |
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| Research Field |
Bacteriology (including Mycology)
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| Research Institution | HIROSHIMA UNIVERSITY |
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Principal Investigator |
YOSHIDA Kazuo HIROSHIMA UNIVERSITY Fac.Science, Biological Science, Professor, 理学部, 教授 (70022720)
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| Co-Investigator(Kenkyū-buntansha) |
KATOH A 米国マーシー病院放射線病理癌部門, 部長
NISHIKAWA Masanobu Okayama Biological Institute, Research Scientist, 技師 (60262812)
TAKEGAMI Tsutomu Knazawa Medical University, Assoiate Professor, 総合医学研究所, 助教授 (10113490)
KOTOH Arthur K. Oncopathology Div., Mercy Hospital, Chief
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| Project Period (FY) |
1996 – 1997
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| Keywords | trans-kindom conjugation / conjugative plasmids / oncogenes / phytooncogenes / Ti pIasmids / animal cuIture celIs / enterobacteria / agrobacteria |
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| Research Abstract |
For trans-kingdom conjugation between prokaryotic Escherichia coli and eukaryotic animal cultured cells, we have constructed a novel plasmid, pBASGreen which consists of oriT,SV40-ori/promoter, pBR322-oriV,Ap, Neo, and GFP (green fluorescent protein) genes. The plasmid was successfully introduced into animal cultured cells, COS1 and NIH3T3 then typical green fluorescence due to the GFP reporter gene was clearly detected in the transformant celIs. However, E.coli donor celIs were unfortunately contaminant and toxic against cultured celIs. To overcome the problems, anucleated and non-dividing celIs of E.coli mini-cell mutant P678-54 was verified to be useful because it is less toxic and keeps the activity of trans-kingdom conjugation. On the basis of conjugal oncogene transfer, we also carried out oncogenes survey in enterobacteria by using Southern blotting. On the other hand, phytopathogenic bacterium Agrobacterium tumefaciens harbors a giant Ti (tumor inducing) plasmid and transfers it's T-DNA into dicots by conjugation-like process. The infected plants subsequently have tumors induced by oncogenes on T-DNA.In order to know the relationship between T-DNA transfer and trans-kingdom conjugation on the molecular basis, we analyzed the entire genome of Ti plasmid. The complete sequencing and analyzing of 203,148bp clearly indicate the independence of conjugation and T-DNA transfer system. We also found that A.tumefaciens is less toxic against animal cultured cells as well as plant celIs. The above results offer important clues on trans-kingdom conjugation between bacterial and animal celIs.
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