Construction and evaluation for the multiplex typing system useful for highly degraded mixed stain specimens

一种新商业发布的试剂盒“Penta BEC多路试剂盒”评估了其在日本人群中的实用性。这些位点上的等位基因频率是从300个不相关的日本人身上计算出来的。在三个位点进行三次检验，均未观察到与Hardy-Weiberg平衡（HWE）的显著偏差。3个位点的辨别力（PD）均大于0.90，累计PD和PE（排除力）分别为0.99991和0.96531。计算了三个位点上的口吃峰值百分比，这些五核苷酸重复STR的峰值百分比低于市售试剂盒中通常使用的四核苷酸重复的峰值百分比。结巴率与等位基因重复数呈正相关，几乎全部分布在每个等位基因的平均±3 SD范围内。因此，每个等位基因的平均值±3 SD可以作为区分混合染色样品产生的口吃峰和次要峰的标准。我们已经基本完成了五核苷酸重复STR基因座（Penta、E、B、D和D10S2325）四重体的构建，其中引物组是新设计的，但还没有完成一些等位基因阶梯标记来确认序列。作为系统评价的一部分，计算了口吃峰值百分比。这些位点的等位基因频率是从日本三个地区人群（秋田、名古屋和大分）中计算出来的。否。在所有位点进行三次测试，观察到与HWE的显著偏差。3个基因座的杂合度和PD值分别大于0.77和0.92。3个位点的等位基因频率两两比较，秋田与大分间2个位点、秋田与名古屋间1个位点、名古屋与大分间1个位点的等位基因频率分布差异有统计学意义（P<0.05）。

A newly commercially released kit "Penta BEC Multiplex kit" was evaluated for its usefulness in a Japanese population. The allele frequencies at those loci were calculated from 300 unrelated Japanese. No significant deviations from Hardy-Weiberg equilibrium(HWE) were observed using three tests at the three loci. The power of discrimination(PD) values for all the three loci were more than 0.90,and the accumulated PD and PE(power of exclusion) were 0.99991 and 0.96531,respectively in this multiplex system. The stutter peak percentages at the three loci were also calculated, those of these pentanucleotide repeat STR were lower than those of tetrranucleotide repeat used generally included in a commercially released kit. Furthermore, the stutter percentages showed the positive correlation with the repeat numbers of the alleles, almost all of them distributed in the range of the mean±3 SD at each allele. Therefore, it was suggested that t the mean±3 SD at each allele could de a standard to distinguish between the stutter peak and the minor peak originated fom a mixed stain sample.We have almost completed to construct a novel quadruplex for pentanucleotide repeat STR loci (Penta, E, B and D, and D10S2325), in which the primer sets were newly designed, but a few of the allelic ladder markers was not finished to confirm the sequences. As a part of the evaluation of the system, the stutter peak percentages were calculated. The allele frequencies at those loci were calculated from three regional Japanese populations (Akita, Nagoya and Oita). No.significant deviations from HWE were observed using three tests at the all loci. The observed heterozygosity and PD values for all the three loci in the three populations were more than 0.77 and 0.92,respectively. Compared pairwisely among the allele frequency distributions of the three loci, two loci between Akita and Oita, one locus between Akita and Nagoya, and between Nagoya-Oita were significantly different (P<0.05).