| Project/Area Number |
12670403
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Legal medicine
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| Research Institution | Iwate Medical University |
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Principal Investigator |
NAKAYASHIKI Nori Iwate Medical University, School of Medicine, Department of Legal Medicine, Assistant Professor, 医学部, 講師 (10146029)
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| Co-Investigator(Kenkyū-buntansha) |
KUMAGAI Reiko Iwate Medical University, School of Medicine, Department of Legal Medicine, Research Associate, 医学部, 助手 (30118294)
KANETAKE Jun Iwate Medical University, School of Medicine, Department of Legal Medicine, Research Associate, 医学部, 助手 (90326661)
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| Project Period (FY) |
2000 – 2001
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| Project Status |
Completed (Fiscal Year 2001)
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| Budget Amount *help |
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 2001: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 2000: ¥1,600,000 (Direct Cost: ¥1,600,000)
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| Keywords | polymorphism / genomic imprinting / DNA methylation / Legal Medicine / 多型 / メチレーション |
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| Research Abstract |
Regions containing SNPs in H19 (maternal expression) and IGF2 gene (paternal expression), which are involved in an imprinting domain on 11p15.5, were investigated concerning existence of diversity on methylation status and selective detection of parental allele in DNA from normal peripheral blood.
At first, we examined known SNPs in H19 and IGF2 gene in order to determine detection methods and find allele distributions. In addition, a haplotype polymorphism composed of three SNPs close to H19 SNP, named H19FR, was analyzed by PCR-SSCP, ARMS PCR and PCR-CDGE, and three major haplotypes with allele frequencies of 0.5177, 0.3409 and 0.1414 were observed.
After treatment of genomic DNA with bisulfite reagent, a methylated allele of H19FR with heterozygous genotype was selectively amplified by methylation-specific PCR. However, polymorphic methylation status in CpGs was not observed in this study.
As another strategy for recognizing parental origin, genomic DNA was treated with methylation-sensitive restriction enzyme and followed by H19FR or H19 SNP typing. As a result, DNA sample having heterozygous genotype in either system showed a single allele derived from methylated paternal origin. The IGF2 SNP revealed, however, only 27 % of maternal allele after the same procedure for H19 SNPs. It is assumable that this method applied for SNPs in upstream region of H19 will be a useful tool for forensic examination to discriminate parental origin.
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