Density separation is a process routinely used to segregate minerals, organic matter, and even microplastics, from soils and sediments. Here we apply density separation to archaeological bone powders before DNA extraction to increase endogenous DNA recovery relative to a standard control extraction of the same powders. Using nontoxic heavy liquid solutions, we separated powders from the petrous bones of 10 individuals of similar archaeological preservation into eight density intervals (2.15 to 2.45 g/cm³, in 0.05 increments). We found that the 2.30 to 2.35 g/cm³ and 2.35 to 2.40 g/cm³ intervals yielded up to 5.28-fold more endogenous unique DNA than the corresponding standard extraction (and up to 8.53-fold before duplicate read removal), while maintaining signals of ancient DNA authenticity and not reducing library complexity. Although small 0.05 g/cm³ intervals may maximally optimize yields, a single separation to remove materials with a density above 2.40 g/cm³ yielded up to 2.57-fold more endogenous DNA on average, which enables the simultaneous separation of samples that vary in preservation or in the type of material analyzed. While requiring no new ancient DNA laboratory equipment and fewer than 30 min of extra laboratory work, the implementation of density separation before DNA extraction can substantially boost endogenous DNA yields without decreasing library complexity. Although subsequent studies are required, we present theoretical and practical foundations that may prove useful when applied to other ancient DNA substrates such as teeth, other bones, and sediments.

中文翻译：

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岩骨粉末的密度分离可优化古 DNA 产量

密度分离是一种通常用于从土壤和沉积物中分离矿物质、有机物甚至微塑料的过程。在这里，我们在 DNA 提取之前对考古骨粉进行密度分离，以相对于相同粉末的标准对照提取提高内源 DNA 回收率。使用无毒的重液体溶液，我们将来自 10 个类似考古保存个体的岩骨中的粉末分离成八个密度区间（2.15 至 2.45 g/cm 3 ，增量为 0.05 ^）。我们发现，2.30 至 2.35 g/cm ³和 2.35 至 2.40 g/cm ³间隔产生的内源独特 DNA 比相应的标准提取多 5.28 倍（在重复读取去除之前最多 8.53 倍），同时保持古代 DNA 真实性的信号，并且不会降低文库的复杂性。尽管较小的 0.05 g/cm ³间隔可以最大程度地优化产量，但去除密度高于 2.40 g/cm ³的材料的单次分离平均产生高达 2.57 倍的内源 DNA，这使得能够同时分离不同密度的样品。保存或分析材料的类型。虽然不需要新的古代 DNA 实验室设备和不到 30 分钟的额外实验室工作，但在 DNA 提取之前实施密度分离可以显着提高内源 DNA 产量，而不会降低文库复杂性。尽管还需要后续研究，但我们提出的理论和实践基础在应用于其他古代 DNA 基质（例如牙齿、其他骨骼和沉积物）时可能会被证明是有用的。