The efficient extraction of DNA from challenging samples, such as bones, is critical for the success of downstream genotyping analysis in molecular genetic disciplines. Even though the ancient DNA community has developed several protocols targeting small DNA fragments that are typically present in decomposed or old specimens, only recently forensic geneticists have started to adopt those protocols. Here, we compare an ancient DNA extraction protocol (Dabney) with a bone extraction method (Loreille) typically used in forensics. Real-time quantitative PCR and forensically representative typing methods including fragment size analysis and sequencing were used to assess protocol performance. We used four bone samples of different age in replicates to study the effects of both extraction methods. Our results confirm Loreille’s overall increased gain of DNA when enough tissue is available and Dabney’s improved efficiency for retrieving shorter DNA fragments that is beneficial when highly degraded DNA is present. The results suggest that the choice of extraction method needs to be based on available sample, degradation state, and targeted genotyping method. We modified the Dabney protocol by pooling parallel lysates prior to purification to study gain and performance in single tube typing assays and found that up to six parallel lysates lead to an almost linear gain of extracted DNA. These data are promising for further forensic investigations as the adapted Dabney protocol combines increased sensitivity for degraded DNA with necessary total DNA amount for forensic applications.

中文翻译：

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使用不同年龄的骨样本评估为法医和古代DNA应用开发的DNA提取方法的评估

从具有挑战性的样品（例如骨头）中有效提取DNA对于分子遗传学领域下游基因分型分析的成功至关重要。即使古代DNA社区已经开发了几种针对小分子DNA片段的方案，这些片段通常存在于已分解或旧的标本中，但直到最近，法医遗传学家才开始采用这些方案。在这里，我们将古老的DNA提取方案（Dabney）与通常用于法医的骨提取方法（Loreille）进行了比较。实时定量PCR和法医代表性打字方法（包括片段大小分析和测序）用于评估方案性能。我们重复使用了四个不同年龄的骨骼样品，以研究两种提取方法的效果。我们的结果证实，当有足够的组织可用时，Loreille的DNA总体上增加的增益，而Dabney的检索较短的DNA片段的效率得到了提高，这在存在高度降解的DNA时是有益的。结果表明，提取方法的选择应基于可用样品，降解状态和目标基因分型方法。我们通过在纯化前合并平行裂解物以研究单管分型测定法中的增益和性能来修改Dabney方案，发现最多六个平行裂解物会导致提取的DNA几乎呈线性增益。这些数据有望用于进一步的法医研究，因为经过修改的Dabney方案将对降解的DNA的敏感性提高与法医应用所需的总DNA量结合在一起。