The selection of new microsatellite loci suitable for fortune forensic exam ination and the establishment of its wide use

在本研究项目为期两年的时间里，我们使用一套用于连锁分析的STR引物集MapPair 8A(Research Genetics Inc.)对人类基因组中168个STR基因座的平均间距进行了分析。这套引物包括168对引物，其中每对中的一对用三基色荧光染料(FAM、HEX、Tet)中的任何一种标记。用基因分析仪310对DNA样品进行扩增，扩增产物按照8A版多重表型，并根据测序结果对每个样品进行分型。我们已经用32个日本人的DNA样本完成了全部168个基因座的分析。从杂合度大于0.8的20个STR基因座中筛选出12个STR基因座，这些基因座的序列结构比较简单。本研究对这些基因座的序列进行了较准确的确认，并对其进行了较小的修正。我们为新系统设计了每个基因座上的每一对引物。一个系统已经完成，另一个几乎已经完成。此外，我们还从诺丁汉大学Armour教授分离的大约20个微卫星基因座中筛选出杂合度较高且没有不规则等位基因的3个STR基因座(wglc5、wgld1、pq2h2)，建立了一个新的复合体系。在第8届日本DNA多态研究学会年会上公布了包括在法医领域的实用性在内的结果。这样，利用15个适合于日语法医鉴定的基因座的新的微卫星分型系统基本完成。

Within the term of this research project (2 years), we analyzed 168 STR loci spacing at 25 cM on average in human genome using a STR primer set for linkage analysis, a MapPair 8A (Research Genetics Inc.). This primer set includes 168 primer pairs in which one of each pair is labeled with either of the tricolor fluorescent dyes (FAM, HEX, TET). DNA samples were amplified and the PCR products were electrophoresed according to the Version 8A multiplex table with the Genetic Analyzer 310, and each sample was typed from sizing results. We have finished to analyze all 168 loci with 32 Japanese DNA samples. Out of 20 STR loci showing heterozygosities of higher than 0.8 twelve STR loci were selected because their sequence structures were simple based on the databases, GDB etc. The sequences of those loci were confirmed accurately in this study with minor corrections.We tried to construct 2 multiplex systems with each 6 STR loci using the tricolor fluorescent dyes. We newly designed each primer set at each locus for the new systems. One system was completed and another has almost been completed. Moreover, we have also devised a mew multiplex system using 3 STR loci (wglc5, wgld1, pq2h2) with comparatively high heterozygosity and without irregular allele selected from approximately 20 microsatellite loci which Prof. Armour in Nottingham University had isolated. The results including the usefulness in the forensic field were presented at the 8th annual meeting of the Japanese Society for DNA polymorphism Research.In this way, the new microsatellite typing systems using 15 loci appropriate to forensic examination in Japanese has almost been completed.