The development of a new HLA-DNA typing method using DNA direct sequencing.

使用 DNA 直接测序开发新的 HLA-DNA 分型方法。

• 批准号: 04557031
• 负责人: KATSUMATA Yoshinao
• 金额: $ 3.78万
• 依托单位: Nagoya University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Developmental Scientific Research (B)
• 财政年份: 1992
• 资助国家: 日本
• 起止时间: 1992 至 1993
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-04557031/
• 关键词: HLA; PCR; Direct Sequence; Forensic Specimens; 法医鑑識; 親子鑑定; 犯罪捜査; 白血球型; 血液型; 遺伝子増幅; 塩基配列直接解読; 蛍光標識プローブ

Highly polymorphic human leukocyte antigen (HLA), which is related to cellular immunoreaction, is useful for personal identification in the field of forensic medicine. To date, HLA class II loci are typed by restriction enzymes or sequence-specific oligonucleotide probes after being amplified by the polymerase chain reaction (PCR). Such procedures are usually complicated and time consuming. Therefore, direct sequencing of the amplified DNA could provide a rapid typing method. However, forensic specimens such as bloodstains, seminal stains, saliva stains and hairs usually contains contaminants which may inhibit PCR reaction. In this study, we tried to analyze the nature of the contaminants, and to minimize the effect of those substances. In hair, we found that melanin is a potent inhibitor for the PCR reaction, and are now trying to minimize the effect of this substances. In bloodstains and saliva stains, we tried to control the inhibition of the contaminated substances, and finally made an improved procedure of DNA purification and typing.So, we started to read the sequence of HLA-DQB1 gene with DNA sequencer.

高度多态性的人类白细胞抗原（HLA）与细胞免疫反应有关，可用于法医学领域的个人身份识别。迄今为止，HLA II 类基因座在通过聚合酶链式反应 (PCR) 扩增后，通过限制性内切酶或序列特异性寡核苷酸探针进行分型。此类程序通常复杂且耗时。因此，对扩增的 DNA 进行直接测序可以提供一种快速分型方法。然而，法医标本如血迹、精液、唾液、毛发等通常含有污染物，可能会抑制PCR反应。在这项研究中，我们试图分析污染物的性质，并尽量减少这些物质的影响。在头发中，我们发现黑色素是 PCR 反应的有效抑制剂，现在正试图尽量减少这种物质的影响。在血迹和唾液污渍中，我们试图控制对污染物质的抑制，最终改进了DNA纯化和分型的程序。于是，我们开始用DNA测序仪读取HLA-DQB1基因的序列。

项目成果

Rieko Uchihi, Keiji Tamaki, Toshinori Kojima, Toshimichi Yamamoto and Yoshinao Katsumata: "Deoxytibonucleic acid (DNA) typing of human leukocyte antigen (HLA)-DQA1 from single hairs in Japanese." Journal of Forensic Sciences. 37(3). 853-859 (1992)

Rieko Uchihi、Keiji Tamaki、Toshinori Kojima、Toshimichi Yamamoto 和 Yoshinao Katsumata：“日本单根毛发中人类白细胞抗原 (HLA)-DQA1 的脱氧核糖核酸 (DNA) 分型。”

Okajima,H.et al.: "Amplification of HLA-DQA1 gene from bloodstains by polymerase chain reaction." Japanese Journal of Legal Medicine. 47(1). 6-12 (1993)

Okajima,H.et al.：“通过聚合酶链式反应从血迹中扩增 HLA-DQA1 基因。”

玉木 敬二: "MUR-PCR法を用いた日本人におけるDIS8(MS32)のタイピングとアリルの解析" 日本法医学雑誌. 46. 474-482 (1992)

Keiji Tamaki：“使用 MUR-PCR 方法对日语中的 DIS8 (MS32) 进行分型和等位基因分析”《日本法医学杂志》46. 474-482 (1992)。

Uchihi,R.et al.: "Deoxyribonucleic acid(DNA)typing of human leukocyte antigen(HLA)-DQA1 from single hairs in Japanese." Journal of Forensic Sciences. 37(3). 853-859 (1992)

Uchihi,R.et al.：“从日本人的单根毛发中对人类白细胞抗原 (HLA)-DQA1 进行脱氧核糖核酸 (DNA) 分型。”

玉木敬二ら: "医学におけるバイオテクノロジー(17)-DNAによる個人識別法-." 現代医学. 40(1). 169-174 (1992)

Keiji Tamaki 等人：“医学生物技术 (17) - 使用 DNA 的个人识别方法。” 169-174 (1992)。