Novel methodology for distinguishing between ancient and contaminating DNA in human archaeological remains

自20世纪80年代末以来，人们已经知道DNA有时保存在人类和其他动物的骨骼和其他遗骸中，在某些情况下，这种“古老”的DNA可以从长达50，000年的标本中提取出来。对古代DNA的研究对动物学产生了巨大的影响，使研究人员能够获得猛犸象和洞熊等灭绝物种的DNA序列，从而对这些动物及其现存亲属之间的进化关系有了新的发现，并为这些物种未能在过去5万年的气候变化中生存下来的原因提供了见解。古DNA可能在考古学中产生同样巨大的影响，例如通过追踪人类骨骼群体之间的家庭关系以及绘制史前人类人口的迁移图。可悲的是，这种潜力尚未实现，原因很简单，骨头等标本很容易被现代人类DNA污染，而且很难将这种污染与标本中真正的古代DNA区分开来。许多研究人员，包括我们，已经提出了解决这个问题的方案，但这些“解决方案”都没有完全成功。到目前为止，我们最好的尝试是基于这样一个事实，即古代DNA在化学上不同于现代DNA，因为古代分子已经部分降解。因此，鉴定骨骼提取物中降解的分子应该可以精确地确定哪些是古老的DNA。但有一个复杂的问题：事实证明，即使是污染的DNA也会在一定程度上降解，因为通常它是在挖掘过程中沉积在标本上的，而这可能是在标本中的DNA被检查之前的几个月或几年，有足够的时间让污染的DNA开始降解。因此，鉴定古代DNA变得更加困难，因为我们不能简单地鉴定哪些分子被降解，而必须区分降解最严重的分子（因此可能是古代DNA）和仅轻微降解的分子（因此可能是现代污染物）。我们尝试用研究古代DNA的标准方法来做这件事，取得了一些成功，但这些方法并不真正适合这种类型的分析。在这个项目中，我们计划测试一种新的方法，称为单引物延伸（dac），这是最近开发的一种工具，用于从灭绝的物种，如恐鸟和塔斯马尼亚狼获得更准确的DNA序列。与标准的古DNA方法不同，dac确实可以准确比较不同DNA分子的损伤量，因此我们相信dac将使我们能够区分人类标本中真正的古DNA和污染物。因此，该项目的目标是将DAC应用于人类骨骼中的DNA分析，看看我们的期望是否得到满足。

Since the late 1980s it has been known that DNA is sometimes preserved in the bones and other remains of humans and other animals, and that in some cases this 'ancient' DNA can be retrieved from specimens up to 50,000 years in age. The study of ancient DNA has had a huge impact in zoology, enabling researchers to obtain DNA sequences from extinct species such as mammoths and cave bears, leading to new discoveries about the evolutionary relationships between these animals and their living relatives, and providing insights into the reasons why these species failed to survive the climate changes of the last 50,000 years. Potentially, ancient DNA could have an equally great impact in archaeology, for example by enabling the family relationships between groups of human skeletons to be traced and by charting the migrations of prehistoric human populations. Sadly, this potential has not been realized, for the simple reason that specimens such as bones are easily contaminated with modern human DNA by handling, and it is difficult to distinguish this contamination from the genuine ancient DNA in a specimen. A number of researchers, including us, have suggested solutions to this problem, but none of these 'solutions' have been completely successful. Our best attempt so far is based on the fact that ancient DNA is chemically different from modern DNA because the ancient molecules have become partially degraded. Identifying the degraded molecules in a bone extract should therefore pinpoint which ones are ancient DNA. But there is a complication: it turns out that even the contaminating DNA is degraded to a certain extent, because usually it is deposited on a specimen during excavation, and this can be months or years before the DNA in the specimen is examined, enough time for the contaminating DNA to begin to degrade. Identifying the ancient DNA therefore becomes more difficult because, rather than simply identifying which molecules are degraded, we must distinguish the ones which are most degraded (and which could therefore be ancient DNA) from the ones that are only slightly degraded (and are therefore likely to be modern contaminants). We have tried to do this with the standard methods used to study ancient DNA, with some success, but these methods are not really suitable for this type of analysis. In this project we plan to test a new method, called single primer extension (SPEX), which was recently developed as a tool for obtaining more accurate DNA sequences from extinct species such as the moa and Tasmanian wolf. Unlike the standard ancient DNA methods, SPEX does allow accurate comparisons of the amounts of damage in different DNA molecules, and we therefore believe that SPEX will enable us to distinguish the genuine ancient DNA in a human specimen from the contaminants. The objective of the project is therefore to apply SPEX to analysis of the DNA in a human bone, to see if our expectations are fulfilled.