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Proboscidean DNA from Museum and Fossil Specimens: An Assessment of Ancient DNA Extraction and Amplification Techniques

机译:来自博物馆和化石标本的长苞亚胺DNa:古DNa提取和扩增技术的评估

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摘要

Applications of reliable DNA extraction and amplification techniques to postmortem samples are critical to ancient DNA research. Commonly used methods for isolating DNA from ancient material were tested and compared using both soft tissue and bones from fossil and contemporary museum proboscideans. DNAs isolated using three principal methods served as templates in subsequent PCR amplifications, and the PCR products were directly sequenced. Authentication of the ancient origin of obtained nucleotide sequences was established by demonstrating reproducibility under a blind testing system and by phylogenetic analysis. Our results indicate that ancient samples may respond differently to extraction buffers or purification procedures, and no single method was universally successful. A CTAB buffer method, modified from plant DNA extraction protocols, was found to have the highest success rate. Nested PCR was shown to be a reliable approach to amplify ancient DNA templates that failed in primary amplification.
机译:可靠的DNA提取和扩增技术对后期样本的应用对古代DNA研究至关重要。测试了来自古代材料的DNA的常用方法,并使用来自化石和当代博物馆的软组织和骨骼进行比较。使用三种主要方法分离的DNAS用作随后的PCR扩增中的模板,并直接测序PCR产物。通过在盲检测系统和系统发育分析下证明再现性,建立了所获得的核苷酸序列的古代起源的认证。我们的结果表明,古代样品可能对提取缓冲液或净化程序进行不同的反应,并且没有单一方法普遍成功。发现从植物DNA提取方案修饰的CTAB缓冲方法具有最高的成功率。嵌套的PCR被证明是一种可靠的方法,用于扩增古代扩增失败的古代DNA模板。

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