1993 Fiscal Year Final Research Report Summary
Studies on genetically engineered bacterium and its introduction into the rumen
| Project/Area Number |
03454100
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
畜産化学
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| Research Institution | Mie University |
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Principal Investigator |
HOSHINO Sadao Mie Univ.Fac.Bioresouces Prof., 生物資源学部, 教授 (90024546)
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| Co-Investigator(Kenkyū-buntansha) |
KOBAYASHI Yasuo Mie Univ.Fac.Bioresources Assis.Prof., 生物資源学部, 講師 (50153648)
WAKITA Masaaki Mie Univ.Fac.Bioresources Associ.Prof., 生物資源学部, 助教授 (40024575)
SAKKA Kazuo Mie Univ.Fac.Bioresources Associ.Prof., 生物資源学部, 助教授 (20154031)
OHMIYA Kunio Mie Univ.Fac.Bioresources Prof., 生物資源学部, 教授 (60023488)
SHIMADA Kyo Mie Univ.Fac.Bioresources Prof., 生物資源学部, 教授 (20024549)
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| Project Period (FY) |
1991 – 1993
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| Keywords | Rumen Bacteria / Fiber Degradation / Ruminococcus albus / Transformation / Conjugal Transfer / Antibiotic Resistance / Plasmid / Goat |
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| Research Abstract |
The present experiment aimed at enhancing fiber-degrading activity of rumen bacteria through a recombinant DNA technique and at successful establishment of the recombinant in the rumen. Attempts to transform rumen bacteria and to inoculate the transformant into the rumen have benn made and the results obtained are as follows : 1)Of 9 plasmids tested, only pRRI207 transformed Ruminococcus albus by electroporation. However, the results were not reproducible. 2)Sequencing of a cryptic plasmid from Butyrivibrio fibrisolvens allowed the replication region to be identified for construction of cloning vector for the host B.fibrisolvens. 3)Suitable markers have been explored to allow easy screening of transformant in the rumen. Genes coding for resistance to at least 3 antibiotics (Rif, Str and Erm) were chosen as the markers. 4)Filter mating procedure was established for a successful conjugal transfer of pAMbeta1 into R.albus. Rif and Str were given to this transconjugant through a spontaneous mutation. 5)The transconjugant resistant to 3 antibiotics was inoculated into goat rumen via a fistulae and ruminal sample was successively taken to determine ruminal level of the transformed R.albus. Untill 37hr after the inoculation the transformant was detected at a level of 10^3/ml of ruminal fluid, though none was found after then. Possible excuses are death and outflow of the transformant, and/or loss of the plasmid from the host. Positive manipulation of bacterial ability to adhere to feed particle as well as more stable vector within a host would be necessary for ecological establishment of engineered bacterium in the rumen.
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