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应用DNA复合条形码技术研究太白山中小型土壤动物多样性

发布时间:2018-08-22 07:40
【摘要】:物种的正确鉴定是所有生物学研究的基础,基于形态学的物种鉴定需要具有丰富经验的分类学家,且通常需要大量的时间,并且表型可塑性和隐存种的存在以及一些物种不同时期的形态学差异会造成错误的鉴定。分类学家的不断减少和基于形态学的物种鉴定的局限性表明我们需要新的方法来完成分类鉴定。2003年,Hebert等提出了DNA条形码的概念。DNA条形码技术的出现弥补了传统分类学存在的一些不足,为分类学的研究提供了重要的帮助。但是,DNA条形码技术只能鉴定单个的生物体,面对混合样本和环境样本时就显现出了其乏力的一面。混合样本和环境样本通常包含了大量的、分类范围广泛的生物个体,分离这些生物个体并一一测定其条形码序列是一项极大的挑战。新一代测序技术的出现与持续发展解决了DNA条形码技术只能鉴定单个生物体的问题。通过新一代测序技术可以获得DNA样品中每一条DNA分子的序列,因此,我们可以提取混合的生物个体的总DNA,选择合适的分子标记进行PCR扩增,然后使用新一代测序技术对PCR产物进行测序,将测得的序列与合适的数据库进行比对,从而获得混合样本或环境样本中包含的分类学信息。这种结合了DNA条形码技术和新一代测序技术的方法就称为DNA复合条形码(DNAmetabarcoding)技术。目前,对于我国中西部地区土壤动物的研究较少。本研究于2014年6月上旬在太白山自然保护区及周围选取高山草甸、针叶林、落叶小叶林、落叶阔叶林和农田5种生境进行了取样,使用Tullgren漏斗分离土样中的中小型土壤动物,以18S rRNA基因和COI基因作为分子标记,应用DNA复合条形码技术对分离得到的中小型土壤动物进行了研究。旨在为进一步探讨秦岭土壤动物多样性格局及其影响因素提供基础资料,为秦岭生物多样性的监测与保护提供科学依据。本研究的主要结果如下:(1)共获得土壤动物424种,隶属于3门10纲34目201科322属,其中高山草甸321种,针叶林315种,落叶小叶林314种,落叶阔叶林339种,农田327种。并对分类到种属水平的类群进行了列表。(2)生境的变化会对中小型土壤动物的群落组成造成一定的影响。不同生境的中小型土壤动物群落组成不同,但各生境的共有物种较多,达217种。而各生境的特有物种很少,分别为:高山草甸10种,针叶林7种,落叶小叶林7种,落叶阔叶林13种,农田8种。
[Abstract]:The correct identification of species is the basis of all biological research. Morphological identification of species requires experienced taxonomists and usually takes a lot of time. Moreover, the existence of phenotypic plasticity and cryptic species, as well as the morphological differences of some species at different stages may lead to false identification. The decreasing number of taxonomists and the limitation of species identification based on morphology indicate that we need new methods to complete classification. In 2003, Hebert et al proposed the concept of DNA barcode. Some of the shortcomings of classicology, It provides important help for the study of taxonomy. However, DNA barcode technology can only identify individual organisms, but it shows a weak side in the face of mixed samples and environmental samples. Mixed samples and environmental samples usually contain a large number of biological individuals with a wide range of classification. It is a great challenge to isolate these organisms and determine their barcode sequences one by one. The emergence and sustainable development of new generation sequencing technology solve the problem that DNA barcode can only identify single organism. The sequence of each DNA molecule in a DNA sample can be obtained by a new generation of sequencing techniques. Therefore, we can extract the total DNA of a mixed organism and select the appropriate molecular markers for PCR amplification. Then the PCR products are sequenced by a new generation of sequencing techniques, and the measured sequences are compared with the appropriate database to obtain the taxonomic information contained in the mixed samples or environmental samples. This method, which combines DNA barcode technology and new generation sequencing technology, is called DNA compound barcode (DNAmetabarcoding) technology. At present, there is little research on soil fauna in the central and western regions of China. In this study, five habitats, alpine meadow, coniferous forest, deciduous leaflet forest, deciduous broad-leaved forest and farmland, were sampled in and around Taibai Mountain Nature Reserve in early June 2014. Small and medium-sized soil animals were isolated from soil samples by Tullgren funnel. Using 18s rRNA gene and COI gene as molecular markers, the isolation of small and medium-sized soil animals was studied by using DNA complex barcode technique. The aim of this study was to provide basic data for further study on soil animal diversity pattern and its influencing factors in Qinling Mountains, and to provide scientific basis for monitoring and protection of biodiversity in Qinling Mountains. The main results are as follows: (1) A total of 424 species of soil animals were obtained, belonging to 322 genera of 201 families, 34 orders, 3 phyla, 10 classes, including 321 species of alpine meadow, 315 species of coniferous forest, 314 species of deciduous leaflet forest, 339 species of deciduous broad-leaved forest and 327 species of farmland. At the same time, the taxonomy to the species level was listed. (2) the community composition of small and medium-sized soil fauna would be influenced by the change of habitat. The composition of small and medium soil fauna communities in different habitats is different, but there are 217 species of common species in different habitats. There are 10 species in alpine meadow, 7 species in coniferous forest, 7 species in deciduous leaflet forest, 13 species in deciduous broad-leaved forest and 8 species in farmland.
【学位授予单位】:陕西师范大学
【学位级别】:硕士
【学位授予年份】:2015
【分类号】:S154.5

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