GIARDIA--A MODEL FOR ANCIENT EUKARYOTIC GENOME ANALYSIS

描述(改编自《调查员摘要》)：这是一份提案 为了检查无性线粒体、寄生原生动物的基因组结构， 蓝氏贾第鞭毛虫。蓝氏囊原虫作为基因组模式生物的筛选 分析基于其对人类健康的公认影响，其 相对中等大小的基因组，包含1200万个碱基对 在五条染色体上，它在分子系统发育中的基础位置，以及 缺乏几个突出的细胞器(如线粒体和 过氧体)是大多数真核细胞的特征。调查人员 将从至少两端确定900-1000个碱基对序列 22,500个随机选择的蓝色脚本克隆，包含3-3.5kb的插入片段。 总体而言，这将提供三倍以上的“通行证”或初选 97%的基因组的数据。他们将合并序列数据 上面收集了现有和未来的映射数据，以组合重叠群 兰布里亚的五条染色体中的每一条。初始序列数据将允许 他们将粘粒文库组装成覆盖98%的 贾第虫基因组。调查人员将使用定向策略来构建 完整的实物地图。他们将在现有的基础上使用初级行走策略 粘粒和质粒克隆用于确定完整的双链序列 与基因相对应的编码区及其侧翼序列 在最近分化的真核生物中或在 三个主要的领域(真核生物、细菌和古生物)。这些地区 对应于可变表面基因簇或发育表达 序列(DEST)也将在两条链上进行测序。分子 系统发育技术将被用来评估分子模式 保守基因的进化。G。基因组序列分析。 兰布里亚将与这个建议的IRPG的功能基因组学相结合 作者：弗朗西斯·吉林。这些应用结合在一起可能会产生新的见解 进入关键途径和细胞器的进化，可变的表面蛋白 被寄生虫用来避开宿主防御机制，以及新的基因 导致真核基因组组织的要素。调查人员 估算测序和测绘方案的总直接成本 (包括信息学和系统发育学)转化为不到 0.20美元/个基数。

DESCRIPTION (Adapted from the Investigator's Abstract): This is a proposal to examine the genome structure of the amitochondriate, parasitic protist, Giardia lamblia. The selection of G. lamblia as a model organism for genome analysis is based on its well-recognized impact on human health, its relatively modest sized genome containing 12 million base pairs distributed onto five chromosomes, its basal position in molecular phylogenies, and the lack of several of the prominent organelles, (e.g. mitochondria and peroxisomes) that characterize most eukaryotic cells. The investigators will determine 900-1000 base pair sequences from both ends of at least 22,500 randomly selected blue script clones containing 3-3.5 kb inserts. Collectively this will provide greater than a threefold "pass" or primary data for 97 percent of the genome. They will incorporate the sequence data gathered above with existing and future mapping data to assemble contigs for each of G. lamblia's five chromosomes. The initial sequence data will allow them to assemble the cosmid library into contigs covering 98 percent of Giardia genome. The investigators will use directed strategies to construct complete physical maps. They will use primer walking strategies on existing cosmid and plasmid clones to determine complete double strand sequences for coding regions and their flanking sequences which correspond to genetic elements that are conserved in more recently diverged eukaryotes or among the three primary domains (Eukarya, Bacteria and Archaea). Regions that correspond to variable surface gene clusters or developmentally expressed sequences (DESTs) will also be sequenced on both strands. Molecular phylogenetic techniques will be employed to assess patterns of molecular evolution for conserved genes. Analyses of genomic sequences from G. lamblia will be coupled with the functional genomics of this IRPG proposed by Francis Gillin. Together these applications may yield novel insights into the evolution of key pathways and organelles, variable surface proteins used by parasites to avoid host defense mechanisms, and novel genetic elements that lead to eukaryotic genome organization. The investigators estimate the total direct cost of the sequencing and mapping proposal (including informatics and phylogenetics) translates into less than $0.20/base.