| Project/Area Number |
06670487
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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 |
内科学一般
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| Research Institution | KYUSHU UNIVERSITY |
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Principal Investigator |
HAYASHUI Jun Kyusyu, University, Faculty of Medicine, Associate Professor, 医学部, 助教授 (20150443)
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| Co-Investigator(Kenkyū-buntansha) |
UENO Kumiko Kyushu, University, Faculty of Medicine, Senior Physician, 医学部, 医員
TANI Yoshiki Kyushu, University, Faculty of Medicine, Senior Physician, 医学部, 医員
NAKASHIMA Koya Kyushu, University, Faculty of Medicine, Senior Physician, 医学部, 医員
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| Project Period (FY) |
1994 – 1995
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| Project Status |
Completed (Fiscal Year 1995)
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| Budget Amount *help |
¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1995: ¥500,000 (Direct Cost: ¥500,000)
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| Keywords | hepatits C virus / HCV RNA / genotype / 遺伝子変異 |
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| Research Abstract |
We developed novel method for genotyping of hepatitis C virus (HCV). This method based on the pattern analysis of fluorescence signal obtained from the reaction utilizing polymerase reaction and dideoxy sequencing reaction.
In brief, reaction was carried out using cDNA synthesized from RNA in sera as template, Tth DNA polymerase as enzyme, dCTP-ddCTP mixture as substrate, and oligonucleotides of conserved HCV sequence as primers. Analysis of resulting fluorescence signal patterndisclosed mutations within targeted 150 bps region with over 97% accuracy. The lower limit of mutant detection was approximately 30% of mutant among viruses in the sample.
This method allowed us to perform genotyping accurately, because highly conserved sequence of HCV can be used to circumvent impaired annealing caused by existing sequence variations among HCV genome. We demonstrated with this method that the prevalence of genotype II and IV were 60% and 40%, respectively, in I island where HCV was highly endemic. That contrasted the repoeted prevalence of genotyoe in Japan where prevalence of genotype II and IV were 80% and 10%, respectively. Furthermore, it has been suggested that transmission of HCV between spouse is rare because of the difference between spouse in genotype or signal pattern. Based on the information of primary HCV genome structure obtained from signal pattern analysis, it is suggested that very few inter-and intraclonal diversity is present in HCV core region when compared with other region and that the magnitude of intraclonal diversity has association with clinical feature of HCV carriers.
Further epidemiological studies are in progress utilizing the partial information of HCV genome variation utilizing this mathod.
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