| Project/Area Number |
04454113
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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 |
基礎獣医学
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| Research Institution | Obihiro University of agriculture and Veterinary Medicine |
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Principal Investigator |
SHINAGAWA Morikazu Obihiro Univ., Vet.Med., Professor., 畜産学部, 教授 (00001537)
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| Co-Investigator(Kenkyū-buntansha) |
HORIUCHI Motohiro Obihiro Univ., Vet.Med., Instructor., 畜産学部, 助手 (30219216)
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| Project Period (FY) |
1992 – 1994
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| Project Status |
Completed (Fiscal Year 1994)
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| Budget Amount *help |
¥6,400,000 (Direct Cost: ¥6,400,000)
Fiscal Year 1994: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 1993: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 1992: ¥4,000,000 (Direct Cost: ¥4,000,000)
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| Keywords | Scrapie / PrP gene / Genotype on PrP / Expression of PrP gene / Replication of scrapie agent / cultured cells / 変異PrP遺伝子 / ウシPrP遺伝子 / 培養細胞 |
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| Research Abstract |
1) Infectious amyloid, which is thought to be an etiological agent of scrapie, consists of PrP^<Sc>, an isoform of a host membrane protein, PrP^C, which is highly expressed in the central nervous system (CNS). It supposedly is one cause of high accumulation of the infectious amyloid in the CNS.We have established a L-929 derived cell-line expressing exogenous PrP (L-BPrP3) by introducing bovine PrP cDNA.In this study, scrapie replication was examined in this cell-line and in the L-929 cell-line. We expected that L-BPrP3 supports sheep scrapie, however, the replication of sheep scrapie in this cell-line was of undetectable levels as it was in the L-929 cell-line, and the mouse-adapted scrapie strain replicated poorly in both cell-lines. These facts suggest that the expression of the bovine PrP gene in L-929 cells did not influence the replication of scrapie agent at all. Therefore, we established another cell-line which expresses sheep PrP at a high level in the PC12 cell-line. The repl
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ication of scrapie agent in this cell-line is beig examined. 2) To analyze PrP in cultured cells expressing two different species of PrP and for the use in immunoprecipitation, we prepared some anti-PrP antibodies. One reacts only with mouse PrP,and another only with sheep PrP.Still another one reacts with both PrPs and can be used for immunoprecipitation. We developed a highly sensitive method to detect PrP using these antibodies. 3) As a part of the boving PrP gene analysis, the promoter region of this gene was analyzed and promoter activity was found in the region between the 1st and the 2nd exon in addition to the upstream region of the 1st exon. 4) In order to choose a Sheep PrP gene for construction of a sheep PrP gene expression vector, we analyzed the sheep PrP gene of various sheep DNAs for amino acid polymorphisms of the PrP,and found a new sheep PrP genotype associated with familiar scrapie with short incubation times. We also estimated the relationships between sheep PrP genotypes and the incidence of onset of scrapie in Japan. Less
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