Zhang Zhen, Wang Bao-Jie, Guan Hong-Yu, Pang Hao, Xuan Jin-Feng
Department of Forensic Serology, China Medical University, Shenyang, China.
J Forensic Sci. 2009 Nov;54(6):1304-9. doi: 10.1111/j.1556-4029.2009.01166.x. Epub 2009 Oct 5.
Reducing amplicon sizes has become a major strategy for analyzing degraded DNA typical of forensic samples. However, amplicon sizes in current mini-short tandem repeat-polymerase chain reaction (PCR) and mini-sequencing assays are still not suitable for analysis of severely degraded DNA. In this study, we present a multiplex typing method that couples ligase detection reaction with PCR that can be used to identify single nucleotide polymorphisms and small-scale insertion/deletions in a sample of severely fragmented DNA. This method adopts thermostable ligation for allele discrimination and subsequent PCR for signal enhancement. In this study, four polymorphic loci were used to assess the ability of this technique to discriminate alleles in an artificially degraded sample of DNA with fragment sizes <100 bp. Our results showed clear allelic discrimination of single or multiple loci, suggesting that this method might aid in the analysis of extremely degraded samples in which allelic drop out of larger fragments is observed.
减小扩增子大小已成为分析法医样本中典型降解DNA的主要策略。然而，当前的微型短串联重复序列-聚合酶链反应（PCR）和微型测序分析中的扩增子大小仍不适用于严重降解DNA的分析。在本研究中，我们提出了一种将连接酶检测反应与PCR相结合的多重分型方法，该方法可用于识别严重片段化DNA样本中的单核苷酸多态性和小规模插入/缺失。此方法采用热稳定连接进行等位基因鉴别，随后进行PCR以增强信号。在本研究中，使用四个多态性位点评估该技术在片段大小<100 bp的人工降解DNA样本中鉴别等位基因的能力。我们的结果显示了对单个或多个位点的清晰等位基因鉴别，表明该方法可能有助于分析观察到较大片段等位基因缺失的极度降解样本。
