| Project/Area Number |
12307011
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| Research Category |
Grant-in-Aid for Scientific Research (A)
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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 | Gunma University |
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Principal Investigator |
KISHI Koichiro Gunma University, Graduate School of Medicine, Professor, 医学部, 教授 (30169841)
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| Co-Investigator(Kenkyū-buntansha) |
TAKESHITA Haruo University of Shimane, Faculty of Medicine, Professor, 医学部, 教授 (90292599)
YASUDA Toshihiro University of Fukui, Faculty of Medicine, Professor, 医学部, 教授 (80175645)
KOMINATO Yoshihiko Gunma University, Graduate School of Medicine, Assistant Professor, 医学部, 助教授 (30205512)
KANEKO Yasushi Gunma University, Graduate School of Medicine, Assistant, 医学部, 助手 (10361370)
NAKAJIMA Tamiko Gunma University, Graduate School of Medicine, Assistant, 医学部, 助手 (40008561)
茂木 康一 群馬大学, 医学部, 助手 (80344919)
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| Project Period (FY) |
2000 – 2003
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| Project Status |
Completed (Fiscal Year 2003)
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| Budget Amount *help |
¥39,180,000 (Direct Cost: ¥33,600,000、Indirect Cost: ¥5,580,000)
Fiscal Year 2003: ¥7,540,000 (Direct Cost: ¥5,800,000、Indirect Cost: ¥1,740,000)
Fiscal Year 2002: ¥8,320,000 (Direct Cost: ¥6,400,000、Indirect Cost: ¥1,920,000)
Fiscal Year 2001: ¥8,320,000 (Direct Cost: ¥6,400,000、Indirect Cost: ¥1,920,000)
Fiscal Year 2000: ¥15,000,000 (Direct Cost: ¥15,000,000)
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| Keywords | Apoptosis / Biochemistry / Deoxyribonuclease / Clinical application / Genetic polymorphism / Molecular biology / Human genetics / Genetic marker / モノクローナル抗体 / 脳下垂体 / 尿 / deoxyribonuclease I / 哺乳類細胞発現系 / 抗体 / ソマトスタチン / 遺伝子発現 / ツメガエル / 膵臓 / cDNA cloning / 両生類 |
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| Research Abstract |
1.We have previously found human deoxyribonuclease (DNase) II to be a genetic marker : DNase II levels in human vary depending on whether the individual has the DNASE2^*H or ^*L allele. It was clarified that the A-75G transition in the proximal promoter region causes the allelic difference in the promoter activity of the gene, underlying its genetic polymorphism.
2.We devised a simple DNA extraction procedure suitable for STR typing of urine samples using a commercially available DNA/RNA extraction kit. This method allows urine samples to be kept in both a frozen or aqueous state for forensic use. Furthermore, we devised a procedure that combines a simple extraction method, isoelectric focusing and activity-staining for DNase I typing from aged urine stains. DNase I polymorphism could be considered the first biochemical marker found to be well suited for individualization from small aged urine stains.
3.We confirmed the population genetic basis for DNase I polymorphism in a Japanese population for forensic application. Our examination of DNase I types revealed a decreasing north-to-south gradient in the DNASE1 allele.
4.We discovered a novel variable number of tandem repeat of 56bp unit in intron 4 of human DNase I gene, resulting in elevation of individualization efficiency of DNase.
5.In order to devise the highly sensitive-and specific-immunological typing for DNase I and II polymorphisms, we succeeded in production of each murine monoclonal anti-human DNase I and DNase II with high specificity using newly developed screening procedure for antibodies. Immunoaffinity procedure using these monoclonal antibodies made the purification of human DNases I and II easier, faster and more effective than the conventional procedure. Furthermore, in order to obtain human DNase I as an immunogen, we developed a procedure consisting of transient expression of the enzyme in transfected mammalian cells with DNase I cDNA and purification of the enzyme from the culture medium.
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