Tuschhoff E J, Hutter Carl R, Glor Richard E
Department of Ecology and Evolutionary Biology, University of Arizona, Tucson, AZ, USA.
Department of Ecology and Evolutionary Biology, University of Kansas, Lawrence, KS, USA.
PeerJ. 2020 Feb 13;8:e8369. doi: 10.7717/peerj.8369. eCollection 2020.
Tissue sample databases housed in biodiversity archives represent a vast trove of genetic resources, and these tissues are often destructively subsampled and provided to researchers for DNA extractions and subsequent sequencing. While obtaining a sufficient quantity of DNA for downstream applications is vital for these researchers, it is also important to preserve tissue resources for future use given that the original material is destructively and consumptively sampled with each use. It is therefore necessary to develop standardized tissue subsampling and loaning procedures to ensure that tissues are being used efficiently. In this study, we specifically focus on the efficiency of DNA extraction methods by using anuran liver and muscle tissues maintained at a biodiversity archive. We conducted a series of experiments to test whether current practices involving coarse visual assessments of tissue size are effective, how tissue mass correlates with DNA yield and concentration, and whether the amount of DNA recovered is correlated with sample age. We found that tissue samples between 2 and 8 mg resulted in the most efficient extractions, with tissues at the lower end of this range providing more DNA per unit mass and tissues at the higher end of this range providing more total DNA. Additionally, we found no correlation between tissue age and DNA yield. Because we find that even very small tissue subsamples tend to yield far more DNA than is required by researchers for modern sequencing applications (including whole genome shotgun sequencing), we recommend that biodiversity archives consider dramatically improving sustainable use of their archived material by providing researchers with set quantities of extracted DNA rather than with the subsampled tissues themselves.
生物多样性档案库中的组织样本数据库是大量遗传资源的宝库，这些组织常常被破坏性地进行二次抽样，并提供给研究人员用于DNA提取及后续测序。虽然为下游应用获取足够数量的DNA对这些研究人员至关重要，但鉴于每次使用原始材料时都是破坏性且消耗性的抽样，为未来保存组织资源也同样重要。因此，有必要制定标准化的组织二次抽样和出借程序，以确保组织得到有效利用。在本研究中，我们特别关注利用保存在生物多样性档案库中的无尾两栖类肝脏和肌肉组织进行DNA提取方法的效率。我们进行了一系列实验，以测试当前基于对组织大小进行粗略视觉评估的做法是否有效，组织质量与DNA产量和浓度之间的关系，以及回收的DNA量是否与样本年龄相关。我们发现，2至8毫克的组织样本提取效率最高，在此范围下限的组织每单位质量能提供更多DNA，而在此范围上限的组织能提供更多的总DNA。此外，我们发现组织年龄与DNA产量之间没有相关性。由于我们发现，即使是非常小的组织二次抽样所产生的DNA往往也远远超过研究人员进行现代测序应用（包括全基因组鸟枪法测序）所需的量，我们建议生物多样性档案库考虑通过向研究人员提供一定量的提取DNA而非二次抽样的组织本身，来大幅提高其存档材料的可持续利用。
