DISSERTATION RESEARCH: Temporal Sampling and DNA Metabarcoding to Test the Climate Variability Hypothesis

论文研究：时间采样和 DNA 元条形码来检验气候变异假说

• 批准号: 1502008
• 负责人: Chris Funk
• 金额: $ 1.93万
• 依托单位: Colorado State University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-06-01 至 2018-05-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1502008&HistoricalAwards=false
• 关键词: DISSERTATION; RESEARCH; Temporal; Sampling; DNA

The Climate Variability Hypothesis (CVH) proposes that the range of temperatures species can tolerate increases with the annual range of temperatures they have experienced over evolutionary time. Climate variability increases with latitude and thus it is expected that temperate species will have wider thermal tolerances, higher dispersal ability, and wider distributions than tropical species. Researchers previously asked if temperate mountain stream insects from Colorado have larger elevation range sizes than tropical species from Ecuador; however, range sizes could only be estimated for common species and specimens could only be collected once. To strengthen this work, stream insects will now be sampled over time (temporal sampling) and a DNA approach will be used to identify all species (common and rare). Temporal sampling will allow researchers to track species' elevational positions as they drift from higher to lower elevations over time. Because the flow of water constantly pushes larvae downstream and temperate larvae spend more time in stream, temperate zone larvae are expected to drift farther downstream during development than tropical species. This work will allow researchers to detect previously overlooked rare species. These advances strengthen an already extensive test of the CVH by incorporation of species movement and rare species into estimates of distributions of elevation range sizes. Results of this work will provide information about how natural selection resulting from climate variability influences species dispersal ability and range sizes with general implications for understanding speciation and the evolution of biodiversity. Larval aquatic insects will be collected 6 times over a yearlong period from streams located along transects from ~1500-3500m in the Colorado Rocky Mountains and the Ecuadorian Andes (11 sites at each latitude). On each sampling date at each site, insects will be collected for 5 minutes from all microhabitats within a 100m stream reach using a D-frame net. Samples will be preserved in ethanol and insects will be separated from sediment and detritus. For each sample, all insect tissue will be homogenized and DNA will be extracted and purified using commercially available kits. The animal DNA barcoding gene, Cytochrome c Oxidase subunit I (COI), will be PCR amplified. Full-length DNA barcodes will be generated by shotgun sequencing and assembly of COI amplicons using next-generation sequencing on an Illumina platform. DNA barcodes will be queried against a regional reference library to determine the composition of samples, re-estimate elevation range sizes for taxa, and compare range sizes and dispersal ability among latitudes.

气候变率假说（CVH）提出，物种可以忍受的温度范围随着它们在进化过程中经历的年温度范围的增加而增加。气候变率随纬度增加而增加，因此预计温带物种将比热带物种具有更大的热耐受性、更高的扩散能力和更广泛的分布。研究人员之前曾问过，来自科罗拉多州的温带山溪昆虫是否比来自厄瓜多尔的热带物种海拔范围更大；然而，范围大小只能估计常见的物种和标本只能收集一次。为了加强这项工作，溪流昆虫现在将随着时间的推移进行采样（时间采样），并将使用DNA方法来识别所有物种（常见和稀有）。时间采样将允许研究人员跟踪物种的海拔位置，因为它们随时间从高海拔到低海拔漂移。由于水流不断地将幼虫推向下游，温带幼虫在河流中停留的时间更长，温带幼虫在发育过程中比热带物种漂流得更远。这项工作将使研究人员能够发现以前被忽视的稀有物种。这些进展通过将物种运动和稀有物种纳入海拔范围分布的估计，加强了已经广泛的CVH测试。这项工作的结果将提供关于由气候变化引起的自然选择如何影响物种扩散能力和范围大小的信息，对理解物种形成和生物多样性的进化具有普遍意义。在一年的时间里，将从科罗拉多落基山脉和厄瓜多尔安第斯山脉沿1500-3500米的横断面（每个纬度11个地点）的溪流中收集6次水生昆虫幼虫。在每个采样点的每个采样日，将使用d型网在100米溪流范围内的所有微生境中采集昆虫5分钟。样品将保存在乙醇中，昆虫将与沉积物和碎屑分离。对于每个样本，所有昆虫组织都将被均质化，DNA将被提取并使用市售试剂盒纯化。动物DNA条形码基因细胞色素c氧化酶亚基I （COI）将被PCR扩增。全长DNA条形码将通过霰弹枪测序和在Illumina平台上使用下一代测序技术组装COI扩增子来生成。DNA条形码将在区域参考库中查询，以确定样品的组成，重新估计类群的海拔范围大小，并比较纬度之间的范围大小和扩散能力。