School of Life Science, University of KwaZulu-Natal, Scottsville, Pietermaritzburg 3209, South Africa.
Mol Ecol Resour. 2015 May;15(3):613-8. doi: 10.1111/1755-0998.12331. Epub 2014 Oct 1.
The global DNA barcoding initiative has revolutionized the field of biodiversity research. Such large-scale sequencing projects require the collection of large numbers of specimens, which need to be killed and preserved in a way that is both DNA-friendly and which will keep voucher specimens in good condition for later study. Factors such as time since collection, correct storage (exposure to free water and heat) and DNA extraction protocol are known to play a role in the success of downstream molecular applications. Limited data are available on the most efficient, DNA-friendly protocol for killing. In this study, we evaluate the quality of DNA barcode (cytochrome oxidase I) sequences amplified from DNA extracted from specimens collected using three different killing methods (ethyl acetate, cyanide and freezing). Previous studies have suggested that chemicals, such as ethyl acetate and formaldehyde, degraded DNA and as such may not be appropriate for the collection of insects for DNA-based research. All Lepidoptera collected produced DNA barcodes of good quality, and our study found no clear difference in nucleotide signal strength, probability of incorrect base calling and phylogenetic utility among the three different treatment groups. Our findings suggest that ethyl acetate, cyanide and freezing can all be used to collect specimens for DNA analysis.
全球 DNA 条形码倡议彻底改变了生物多样性研究领域。这种大规模测序项目需要收集大量的标本，这些标本需要以既对 DNA 友好又能保持凭证标本良好状态以供以后研究的方式进行处理和保存。已知采集后时间、正确的储存（避免自由水和热量）以及 DNA 提取方案等因素对下游分子应用的成功起着重要作用。关于最有效、对 DNA 友好的致死方法，目前可用的数据有限。在这项研究中，我们评估了从使用三种不同致死方法（乙酸乙酯、氰化物和冷冻）采集的标本中提取的 DNA 扩增的 DNA 条形码（细胞色素氧化酶 I）序列的质量。先前的研究表明，化学物质，如乙酸乙酯和甲醛，会降解 DNA，因此可能不适合用于基于 DNA 的昆虫收集研究。所有采集的鳞翅目昆虫都产生了高质量的 DNA 条形码，我们的研究发现，在三种不同处理组之间，核苷酸信号强度、错误碱基调用的概率和系统发育效用没有明显差异。我们的研究结果表明，乙酸乙酯、氰化物和冷冻都可用于收集用于 DNA 分析的标本。
