Department of Legal Medicine, Asahikawa Medical University, 2-1-1-1 Midorigaoka-higashi, Asahikawa 078-8510, Japan; Oral and Maxillofacial Surgery, Asahikawa Medical University, 2-1-1-1 Midorigaoka-higashi, Asahikawa 078-8510, Japan.
Mol Cell Probes. 2014 Feb;28(1):13-8. doi: 10.1016/j.mcp.2013.09.002. Epub 2013 Sep 25.
Short insertion/deletion (Indel) polymorphisms of approximately 2-6 bp are useful as biallelic markers for forensic analysis, and the application of Indel genotyping as a supplementary tool would improve human identification accuracy. We examined the allele frequencies of 37 autosomal Indels in the Japanese population and developed a novel dual-color genotyping method for human identification on the basis of universal fluorescent PCR, including the sex-typing amelogenin locus. Target genomic fragment sizes for 38 Indels were 49-143 bp. We analyzed these Indels in 100 Japanese individuals using the M13(-47) sequence as a universal primer. For dual-color genotyping, we designed a novel universal primer with high amplification efficiency and specificity. Using FAM-labeled M13(-47) and HEX-labeled modified M13(-47) primers, fluorescent signals at all loci were clearly distinguished in two independent multiplex PCRs. Average minor allele frequency was 0.39, and accumulated matching probability was 2.12 × 10(-15). Complete profiles were successfully amplified with as little as 0.25 ng of DNA. This method provides robust, sensitive, and cost-effective genotyping for human identification.
短插入/缺失（Indel）多态性约为 2-6 bp，可用作法医分析的双等位基因标记，Indel 基因分型的应用作为补充工具将提高人类识别的准确性。我们研究了日本人群中 37 个常染色体 Indel 的等位基因频率，并基于通用荧光 PCR 开发了一种新的人类鉴定双色基因分型方法，包括性别鉴定的釉原蛋白基因座。38 个 Indel 的目标基因组片段大小为 49-143 bp。我们使用 M13(-47) 序列作为通用引物分析了 100 名日本人中的这些 Indel。对于双色基因分型，我们设计了一种新型的通用引物，具有高扩增效率和特异性。使用 FAM 标记的 M13(-47) 和 HEX 标记的改良 M13(-47) 引物，在两个独立的多重 PCR 中可以清楚地区分所有位点的荧光信号。平均次要等位基因频率为 0.39，累积匹配概率为 2.12×10(-15)。即使使用低至 0.25 ng 的 DNA 也可以成功扩增完整的图谱。该方法为人类鉴定提供了稳健、敏感和具有成本效益的基因分型。
