An Automated DNA Sequencer for Analysis of Gene Structure and Function

本申请要求部分支持自动DNA测序设备的开发。华盛顿大学生物化学系、植物学系、遗传学系和动物学系的7个独立实验室（主要用户）和这些系的另外7个实验室（次要用户）正在参与这项提议，这些实验室对DNA测序的需求很大，但不太广泛。这一应用的推动力来自于这些人的认识，即现有的人工DNA测序方法比使用荧光化合物跟踪DNA片段的自动化方法更慢、工作量更大、成本更高。参与这项申请的所有14名教员都得出结论，需要依靠人工方法进行DNA测序，这对他们的研究进展产生了明显的负面影响。包含本申请的14个项目全部由美国国家科学基金会、美国国立卫生研究院和其他资助机构提供的一项或多项研究资助。主要项目包括：果蝇卵形成过程中细胞迁移所需基因的分析（C. Berg）；哺乳动物metaxin基因家族的结构和功能（P. bernstein）；海洋植物系统分子分析（R. Cattolico）；鸟类MHC II类区的大规模测序（S. Edwards）；不同分类群核RNA聚合酶II两个最大亚基的比较序列分析（B. Hall）；肌酸激酶基因调控分析（S. Hauschka）；昆虫发育的激素调节（L. Riddiford）。次要项目包括：拟南芥（L. Comai）的revolta基因的表征；酿酒酵母微管组织中心成分的鉴定酵母细胞周期的遗传控制（L. Hartwell）；爪蟾和斑马鱼中胚层模式的调控（D. Kimelman）；果蝇发育中的极性起源（H. Ruohola-Baker）；果蝇的异色基因及其位置效应酵母中调节蛋白及其靶基因的进化[E.Young]。我们要求购买一台377型ABI Prism自动DNA测序仪及其相关的计算机软件和一台GeneAmp PCR System 9600。这些仪器将被安置在生物化学系提供的空间里。该设施将以按服务收费的方式运行（成本中心）。预计该设施的80-90%的能力将由参与的实验室使用，剩余的能力将按先到先得的原则提供给华盛顿大学的研究人员。

This application requests partial support for the development of an Automated DNA Sequencing Facility. Seven independent laboratories in the Departments of Biochemistry, Botany, Genetics, and Zoology at the University of Washington, for whom DNA sequencing represents a major research focus (major users), and an additional seven laboratories in these Departments with a significant but less extensive need for DNA sequencing (minor users), are participating in this proposal. The impetus for this application derives from the realization by these individuals that existing methods for manual sequencing of DNA are slower, more work-intensive, and more expensive than are automated methods that employ fluorescent compounds to track DNA fragments. All 14 faculty members involved in this application have concluded that the need to rely on manual methods for DNA sequencing is having a distinctly negative impact on the progress of their research. The 14 projects that comprise this application are all funded by one or more research grants from the National Science Foundation, National Institutes of Health, and other granting agencies. The major projects involve: an analysis of genes required for cell migrations during oogenesis in Drosophila (C. Berg); the structure and function of the mammalian metaxin gene family (P. Bornstein); a molecular analysis of marine plant systems (R. Cattolico); large-scale sequencing of the MHC class II region in birds (S. Edwards); comparative sequence analysis of the two largest subunits of nuclear RNA polymerase II from different taxa (B. Hall); regulatory analyses of muscle creatine kinase genes (S. Hauschka); and hormonal regulation of insect development (L. Riddiford). The minor projects involve: characterization of the revolta gene of Arabidopsis thaliana (L. Comai); identification of components of the microtubule organizing center in S. cerevisiae (T. Davis); genetic control of the yeast cell cycle (L. Hartwell); regulation of mesodermal patterning in Xenopus and zebrafish (D. Kimelman); origin of polarity in Drosophila development (H. Ruohola-Baker); heterochromatic genes and position effects in Drosophila (B. akimoto); and evolution of a regulatory protein and its target gene in yeast (E.Young). We request the purchase of a model 377 ABI Prism Automated DNA Sequencer with associated computer software, and a GeneAmp PCR System 9600. These instruments will be housed in space provided by the Department of Biochemistry. The facility will be run on a fee-for-service basis (cost center). It is anticipated that 80-90% of the capacity of the facility will be utilized by the participating laboratories and that the remaining capacity will be made available to investigators at the University of Washington on a first-come, first-served basis.