Marine Biotech Fellowship: Genetic Population Structure and Gene Flow in Deep-Sea Invertebrates

9212994法国 深海是地球上最大的环境，已知其无脊椎动物群高度多样化。 虽然已作出相当大的努力来衡量物种多样性的格局，但对产生深海动物的进化力量的研究仍然很少。为了估计基因流和种群分化和物种形成的潜力，需要研究地理变异的遗传模式。然而，只有少数这样的研究是在深海中进行的。量化深海物种遗传群体结构的主要障碍之一是难以获得足够数量的适当保存的组织用于遗传研究。大多数深海底栖生物是由小的、分布稀疏的生物组成，在典型的取样过程中，这些生物是死亡或垂死的;这些标本不足以用于等位酶电泳的标准技术。 然而，随着新的分子生物学技术的出现，可以对古老的、干燥的和酒精保存的组织的DNA序列进行鉴定，对深海物种群体遗传学的研究现在是可行的。特别是，从几十次深海取样工作中收集的大量标本提供了大量可供分析的潜在信息。该项目将应用聚合酶链反应技术，确定深海端足类动物酒精保存标本的遗传变异特征，目的是估计海洋内和海洋间地理尺度上的遗传种群结构和基因流动。为了实现这一目标，该项目将：1。用PCR，用从果蝇mtDNA序列构建的通用引物扩增每个群体中15-25个个体的线粒体DNA（mtDNA）的快速进化部分; 2.）检查高变区的扩增片段; 3.）鉴定15-20个碱基对区域，其中大多数变异限于1或2个多态性核苷酸，并为每个等位基因设计等位基因特异性寡核苷酸（阿索）探针; 4.将阿索探针与来自每个群体100-200个个体的扩增的高变区杂交; 5）计算每个群体中的等位基因和基因型频率并评估群体结构。 这一项目的结果将首次详细审查深海盆地和世界各大洋之间的遗传种群结构。此外，该项目有可能展示分子生物学技术对深海生态问题的适用性，并促进进一步利用现有资源解决重要的海洋演变问题。 ***

9212994 France The deep sea is the largest environment of Earth and is known to be highly diverse in its invertebrate fauna. Although considerable effort has been devoted to measuring the patterns of species diversity, the evolutionary forces which have generated the deep- sea fauna remain poorly studied. In order to estimate gene flow and the potential for population differentiation and speciation, studies of the genetic patterns of geographic variation are required. Yet only a handful of such studies have been done in the deep sea. One of the major obstacles to quantifying genetic population structure in deep-sea species has been the difficulty of obtaining sufficient quantities of appropriately preserved tissue for genetic studies. Most of the deep-sea benthos is comprised of small, sparsely distributed organisms which are recovered dead or moribund during typical sampling; these specimens have been inadequate for the standard techniques of allozyme electrophoresis. However, with the advent of new molecular biological techniques which make possible the characterization of DNA sequences from ancient, dried, and alcohol-preserved tissues, studies of the population genetics of deep-sea species are now feasible. In particular, the extensive collections of specimens from dozens of deep-sea sampling efforts provide a wealth of potential information ready to be analyzed. This project will apply the techniques of the polymerase chain reaction (PCR) to characterize genetic variation from alcohol-preserved specimens of deep-sea amphipods, with the goal of estimating genetic population structure and gene flow at intra-ocean and inter-ocean geographic scales. To accomplish this goal, this project will, 1.) with PCR, amplify rapidly evolving portions of the mitochondrial DNA (mtDNA) from 15-25 individuals in each population with universal primers constructed from mtDNA sequences of Drosophila; 2.) inspect amplified segments for hypervariable regions; 3.) identify 15-20 base pair regions where most variation is confined to 1 or 2 polymorphic nucleotides and design allele-specific oligonucleotide (ASO) probes for each allele; 4. hybridize ASO probes to amplified hypervariable regions from 100-200 individuals per population; 5) calculate allele and genotype frequencies in each population and evaluate population structure. The results of this project should provide the first detailed examination of genetic population structure across deep ocean basins and between world oceans. In addition, this project has the potential to demonstrate the applicability of molecular biological techniques to problems in deep-sea ecology and to promote further use of available resources in addressing important evolutionary issues in the oceans. ***