CAA: Technology Transfer: Bringing the Tools of Molecular Biology to an Ecological Genetics Program

摘要苏珊卡利什 小行星9707679 测量自然界中种群内和种群间个体的扩散或运动速率仍然是生态学家和种群遗传学家面临的挑战，因为难以检测个体运动。 这个困难是 对于许多植物物种来说，这一点被夸大了，在这些植物中，花粉或种子的传播阶段很小。然而，我们量化植物种群内和种群间扩散或迁移的能力对于理解许多种群和区域过程至关重要。 特别地，种群或亚种群通过花粉或种子的迁移而连接的程度将决定1）不同种群或亚种群之间的差异程度。 种群共享共同基因的程度; 2）局部适应与区域内其他种群的同化平衡的程度; 3）局部适应发生的程度。 新兴的DNA技术现在使它 可能使用遗传标记以更准确地检测迁移和异交事件。 PI将学习一种强大的生态和群体遗传学新分子技术，即制作和筛选微卫星DNA重复序列和微卫星DNA多态性分析的基因组文库。 微卫星 是短的、串联重复的简单序列，长度为1至6个碱基对，其重复数高度多态。通过聚合酶链反应（PCR）扩增微卫星区域，导致完全外显，孟德尔遗传， 可以通过长度精确识别的共显性标记。由于这些标记是等位的，微卫星数据直接适用于群体遗传模型和分析。 微卫星DNA标记的最大优点之一是其 极端变异性（通常10-20个等位基因/基因座），可用于表征群体结构、确定异交率和通过亲子排除来评估亲子关系。 该奖项将促进微卫星文库生产的所有方面， 克隆、筛选和评分：本质上，就是将这种基于PCR的技术转移到我的项目中。这些技术将从M博士那里学到。阿什利在她的实验室里为几种植物和动物系统开发了微卫星标记。 微卫星标记的分析将促进四个不同领域的当前研究和未来发展：1）自然群体中典型的环境变异导致局部变异的发生。这种当地人口灭绝 应该选择对冲的特征，恢复人口，人口统计学和遗传。 关于局部隔离，我们假设存在于同一物种的其他种群的基质中的植物种群应该 种子传播的经验选择。 与此相反，孤立的人口应该经历选择种子休眠，因为孤立的人口不能接收，也不能输出成功的移民。一个目标是量化种子与 扩散、种子休眠和种群遗传结构。微卫星标记将允许评估的程度，其中的研究种群的Collinsia verna是遗传隔离的实体（和表达种子休眠表型）或通过种子传播（和缺乏种子休眠）交换移民。2)大多数自交进化模型认为，近交衰退的负遗传效应是交配系统进化的关键因素。 然而，一个新的模型表明，环境的变化，禁止传粉者访问花在开花季节的某些部分可能比近亲繁殖的抑郁症更重要的驱动进化和维护的自花授粉机制。 我们 证明了C. Verna具有高水平的异交和雄性和雌性阶段的时间分离，但能够自交。 微卫星标记将使我们能够通过进行亲子分析和花内分析来检验这一模型 异交率

Abstract Susan Kalisz 9707679 CAA Measuring dispersal or the rate of movement of individuals within and among populations in nature remains a challenge for ecologists and population geneticists because of the difficulty in detecting individual movements. This difficulty is exaggerated for many plant species, in which the dispersing stages are small, such as pollen or seed. However, our ability to quantify dispersal or migration within and among plant populations is central to understanding many population and regional processes. In particular, the extent to which populations or sub-populations are connected via migration of pollen or seed will determine 1) the extent to which different populations share genes in common and 2) the extent to which local extinctions are balanced by recolonization from other populations within a region, and 3) the extent to which local adaptation will occur. Emerging DNA technologies now make it possible to use genetic markers to detect migration and outcrossing events with greater accuracy. A powerful new molecular technique for ecological and population genetics, that of making and screening genomic libraries for microsatellite DNA repeats and microsatellite DNA polymorphism analysis will be learned by the PI. Microsatellites are short, tandemly repeated simple sequences, one to six base pairs in length, that are highly polymorphic for repeat number. Amplification of the microsatellite region by polymerase chain reaction (PCR) results in fully penetrant, Mendelian-inherited, codominant markers that can be precisely identified by length. Since these markers are allelic, microsatellite data are directly applicable to population genetic models and analysis. One of the greatest advantages of microsatellite DNA markers is their extreme variability (typically 10-20 alleles/locus) that can be used to characterize population structure, determine outcrossing rates and evaluate parentage by paternity exclusion. This award will facilitate al l facets of microsatellite library production, cloning, screening and scoring: in essence, the transfer of this PCR-based technology to my program. These techniques will be learned from Dr. M. Ashley who has developed microsatellite markers for several plant and animal systems in her laboratory. The analysis of microsatellite markers will enhance current research and future development in four distinct areas: 1) Environmental variance typical in natural populations causes local extinctions to occur. Such local population extinction should select for bet-hedging traits which restore populations, both demographically and genetically. With respect to local extinctions, we hypothesize that plant populations which exist within a matrix of other population of the same species should experience selection for seed dispersal. In contrast, isolated populations should experience selection for seed dormancy, since isolated population cannot receive nor cannot export successful migrants. One goal is to quantify the relationship between seed dispersal, seed dormancy and population genetic structure. Microsatellite markers would allow the assessment of the extent to which the study populations of Collinsia verna are genetically isolated entities (and express seed dormancy phenotypes) or exchange migrants via seed dispersal (and lack seed dormancy). 2) Most models of the evolution of selfing consider the negative genetic effects of inbreeding depression as the critical factors in mating system evolution. However, a new model demonstrates that environmental variance which prohibits pollinators from visiting flowers during some portion of the flowering season can be more important than inbreeding depression in driving the evolution and maintenance of self pollination mechanisms. We have demonstrated that C. verna has high levels of outcrossing and temporal separation of male and female phases but is capable of selfing. Microsatellite markers would allow us to test this model by conductin g paternity analysis and within flower outcrossing rates.