Harper G L, King R A, Dodd C S, Harwood J D, Glen D M, Bruford M W, Symondson W O C
Cardiff School of Biosciences, Cardiff University, PO Box 915, Cardiff, CF10 3TL, UK.
Mol Ecol. 2005 Mar;14(3):819-27. doi: 10.1111/j.1365-294X.2005.02442.x.
DNA-based techniques are providing valuable new approaches to tracking predator-prey interactions. The gut contents of invertebrate predators can be analysed using species-specific primers to amplify prey DNA to confirm trophic links. The problem is that each predator needs to be analysed with primers for the tens of potential prey available at a field site, even though the mean number of species detected in each gut may be as few as one or two. Conducting all these PCRs (polymerase chain reactions) is a lengthy process, and effectively precludes the analysis of the hundreds of predators that might be required for a meaningful ecological study. We report a rapid, more sensitive and practical approach. Multiplex PCRs, incorporating fluorescent markers, were found to be effective at amplifying degraded DNA from predators' guts and could amplify mitochondrial DNA fragments from 10+ species simultaneously without 'drop outs'. The combined PCR products were then separated by size on polyacrylamide gels on an ABI377 sequencer. New primers to detect the remains of aphids, earthworms, weevils and molluscs in the guts of carabid predators were developed and characterized. The multiplex-sequencer approach was then applied to field-caught beetles, some of which contained DNA from as many as four different prey at once. The main prey detected in the beetles proved to be earthworms and molluscs, although aphids and weevils were also consumed. The potential of this system for use in food-web research is discussed.
基于DNA的技术为追踪捕食者与猎物之间的相互作用提供了宝贵的新方法。可以使用物种特异性引物分析无脊椎动物捕食者的肠道内容物，以扩增猎物DNA来确认营养联系。问题在于，即使在每个肠道中检测到的物种平均数量可能只有一两种，但对于野外现场存在的数十种潜在猎物，每个捕食者都需要用相应的引物进行分析。进行所有这些聚合酶链反应（PCR）是一个漫长的过程，实际上排除了对有意义的生态研究可能需要的数百个捕食者进行分析的可能性。我们报告了一种快速、更灵敏且实用的方法。发现结合荧光标记的多重PCR能够有效地扩增捕食者肠道中降解的DNA，并且可以同时从10多种物种中扩增线粒体DNA片段而不会出现“遗漏”。然后，将合并的PCR产物在ABI377测序仪上的聚丙烯酰胺凝胶上按大小进行分离。开发并鉴定了用于检测步甲捕食者肠道中蚜虫、蚯蚓、象鼻虫和软体动物残骸的新引物。然后将多重测序仪方法应用于野外捕获的甲虫，其中一些甲虫同时含有多达四种不同猎物的DNA。在甲虫中检测到的主要猎物是蚯蚓和软体动物，不过蚜虫和象鼻虫也有被食用。本文讨论了该系统在食物网研究中的应用潜力。
