Development on practical examination methods using DNA polymorphisms in the field of Forensic Dentistry

本研究的目的是建立一种实用可靠的方法，用于法医口腔医学中使用高度降解的DNA源检测DNA多态性。在第一年中，我们检查了ST上PC大小的减少是否提供了可重复的结果，以及它是否提高了降解DNA的扩增效率。比较了D12 SG 6、VWA和CD 4等小片段ST基因座扩增的可靠性，并考察了从同一多态性区域产生不同大小PCB产物的引物的构建及其应用于PC时对分型的影响。结果表明，PC大小的减少增加了从降解DNA的扩增效率，但不协调分型的可靠性。我们调查了201个日本人的线粒体DNA控制区的序列多态性，并根据相邻连接分析的结果将165个单倍型分为32个簇。这些特征表明mtDNA多态性可能应用于法医生物学样本的族群估计。我们还调查了日本人群中的10个Y染色体ST多态性，并显示了DYS 390位点的结构变异，从而鉴定了Y-ST多态性的高信息鉴别力。由于ST多态性在降解DNA的PCR中有时会导致错误的结果，我们通过设计引物扩增小片段的PCR产物，扩展了我们对Y-SNPs的研究，使从高度降解的DNA中获得扩增产物成为可能。还检查了常染色体ST多聚体和SNP，以比较降解DNA样品分型的可靠性。Y-SNPs单倍型分析表明，在日本人口的男性个体中存在几个谱系。我们还展示了患者使用的程序和树脂。

The purpose of this study is to establish practicable and reliable methods to detect DNA polyomorphisms using highly degraded DNA sources in Forensic Dentistry. In the first year, we examined if reduction of the PC size on STs gave reproducible results and also if it improved amplification efficiency from degraded DNAs. We compared reliability of amplification of the small-sized ST loci, such as D12SG6, VWA and CD4 and examined influences on typing when the primers producing different-sized PCB products from the same polymorphic region were constructed and applied to the PC. The results indicated that reduction of the PC size increased amplification efficiency from degraded DNAs, but did not coordinate reliability of typing. We investigated sequence polymorphism of mitochondrial DNA control regions from 201 Japanese individuals and classified 165 haplotypes into 32 clusters based on the results of Neighbour Joioning analysis. Each clusters shared specific polymorphic sites, and these characteristics suggested possible application of mtDNA polymorphisms to ethnic group estimation from forensic biological samples. We also investigated 10 Y chromosome ST polymorphisms in the Japanese population and also shoed structural variation in the DYS390 locus, resulting in the identification of highly informative discriminatory power of Y-ST polymorphisms. Beauses ST polymorphisms sometimes gave erroneous results in PC of degraded DNA, we extended our studies on Y-SNPs by designing primers to amplify small PC products, which made it possible to obtain amplification products from highly degraded DNAs. Autosomal ST polymerphisms and SNPs were also examined to compare reliability of typing from degraded DNA samples. Haplotype analysis on Y-SNPs indicated the presence of several lineages in male individuals in the Japanese population. We also showed procedure and resin used by the patients.