Acquisition of Instrumentation for Automated DNA Analyses

我们建议建立一个DNA分析设施，购买几台关键仪器，用于DNA分子的自动分析。所要求的仪器是市售的，代表了其他主要研究型大学许多基因组和DNA测序中心的核心。这种类型的设备对于需要在全基因组范围内进行实验的研究项目是必不可少的。目前在俄勒冈大学有三个这样的研究项目；如果要实现其既定目标，所有这些都绝对需要建立拟议的DNA分析设施。此外，所要求的仪器的采购可能会有显著的辅助效益。通过将自动化方法纳入他们的研究计划，许多其他仪器用户将能够更有效和更经济地完成他们的项目。DNA分析设施的建立也将极大地帮助我们招募对发育遗传学，分子神经科学和分子进化研究感兴趣的新教师。这些都是我们计划在不久的将来雇用新教员的领域。具体来说，需要资金用于：1)集成分离系统AutoGen 740，可以在24小时内制备多达200个病毒DNA，细菌质粒DNA，酵母人工染色体（YAC） DNA或基因组DNA样本；2) Biomek 2000机器人工作站和侧载机，用于克隆和核酸的机器人操作；3) ABI 377 DNA测序仪及进行高通量自动DNA测序和微卫星分析所需的附件。此外，还需要几件小型设备（包括Macintosh PowerPC 8100、两台GeneAmp 9600热循环仪和两台微生物培养液）来装备DNA分析设施。目前，在俄勒冈州没有任何设施可以为拟议的DNA分析设施提供研究支持。这项提案的资金将允许教员充分发展最近启动的项目，这些项目利用复杂（数量）性状遗传分析方面的新兴智力和技术进步，并利用基于图谱的克隆策略来分离和表征主要影响的基因。这些项目都旨在阐明基本生物过程的遗传和分子基础，并涵盖广泛的生物学，从进化和种群遗传学，到发育遗传学，再到行为遗传学。例如，将由这些仪器支持的植物遗传学项目将识别植物基因组中的基因，这些基因控制着单个物种中不同植物繁殖策略的进化，这些物种的个体在地理上被隔离了许多代。狗项目使用与植物项目相同的实验逻辑，但在这种情况下，计划是利用狗品种之间显著的形态和行为差异来深入了解这些差异的遗传和分子基础。斑马鱼的研究工作集中在鉴定对生物体胚胎发育重要的基因，包括神经系统正常发育和功能所需的基因。这些研究人员正在追求的一个重要方法是确定影响有关发育过程的新突变。将这些新突变放置到高质量遗传图谱上的能力将促进基因的克隆，并最终对这些基因功能的分子理解。应该指出的是，本提案中描述的许多项目本质上是协作的。这反映了我们的教师努力结合我们的个人专业知识，以便更快地推进我们感兴趣的领域。随着所需仪器的获得，建立DNA分析设施将代表这些合作努力的重大进展，并将为我们的研究计划的成功提供必要的基础设施。

We propose to establish a DNA analysis facility with the purchase of several key instruments for the automated analysis of DNA molecules. The instrumentation requested is commercially available and represents the core of many genome and DNA sequencing centers at other major research universities. This type of facility is essential for research projects that require experiments to be done on a genome-wide scale. There are currently three such research programs at the University of Oregon; all of these absolutely require the establishment of the proposed DNA analysis facility if they are to achieve their stated goals. In addition, the acquisition of the requested instrumentation is likely to have significant ancillary benefits. Many other users of the instruments will be able to complete their projects more efficiently and economically by incorporating the automated methods into their research programs. The establishment of the DNA analysis facility should also greatly aid our recruitment of new faculty with research interests in developmental genetics, molecular neuroscience and molecular evolution. These are all areas in which we plan to hire new faculty in the near future. Specifically, funds are requested for: 1) an Integrated Separation Systems AutoGen 740 that can prepare up to 200 DNA samples of viral DNA, bacterial plasmid DNA, yeast artificial chromosomal (YAC) DNA or genomic DNA in a 24 hour period; 2) a Biomek 2000 robotic workstation and sideloader for robotic manipulations of clones and nucleic acids; and 3) an ABI 377 DNA sequencer and the necessary accessories to perform high throughput automated DNA sequence and microsatellite analysis. In addition, several small equipment items (including Macintosh PowerPC 8100, two GeneAmp 9600 thermocyclers, and two microfuges) are requested to equip the DNA analysis facility. Presently, there is no facility in the State of Oregon that can provide the research support of the proposed DNA analysis facility. Fundin g this proposal will allow the faculty to fully develop recently initiated projects that utilize emerging intellectual and technical advances in the genetic analysis of complex (quantitative) characters, and that take advantage of map-based cloning strategies to isolate and characterize genes of major effect. These projects all aim to elucidate the genetic and molecular basis of fundamental biological processes, and encompass a broad spectrum of biology, from evolution and population genetics, to developmental genetics, to behavioral genetics. For example, the plant genetics project that will supported by these instruments will identify genes within the plant genome that control the evolution of different strategies of plant reproduction within a single species in which individuals have been geographically isolated for many generations. The dog project uses the same experimental logic being applied in the plant project, but in this case, the plan is to take advantage of the striking morphological and behavioral differences between dog breeds to gain insight into the genetic and molecular basis for these differences. The zebrafish research efforts are concentrated on identifying genes important for embryonic development of the organism, including genes required for the proper development and functioning of the nervous system. An important approach which is being pursued by these investigators is to identify new mutations that effect developmental processes in question. The ability to place these new mutations onto a high quality genetic map will facilitate the cloning of the genes and eventual molecular understanding of how these genes function. It should be noted that many of the projects described in this proposal are collaborative in nature. This reflects the efforts of our faculty to combine our individual expertise in order to advance our fields of interest more rapidly. Establishing the DNA analysis facility with the acquisition of the requested instrumentati on will represent a major advance in these collaborative efforts, and will provide the infrastructure necessary for the success of our research programs.