Expanding the Xenopus ORFeome to genome-scale by de novo cloning of protein-coding gene models

通过蛋白质编码基因模型的从头克隆将爪蟾 ORFeome 扩展到基因组规模

基本信息

批准号：
9279858
负责人：
Michael James Gilchrist
金额：
$ 78.87万
依托单位：
UNIVERSITY OF VIRGINIA
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-05-01 至 2020-02-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9279858
关键词：
Address Animal Model Biochemical Bioinformatics Biological Biological Assay Biological Models Biomedical Research Cell Cycle Cellular biology Cloning Code Collection Communities Complementary DNA Data Databases Developmental Biology Funding Gene Chips Genes Genome Grant Human Initiator Codon Institutes Investments Laboratories Length Marines Messenger RNA Modeling Molecular Biology Neurobiology Open Reading Frames Organism Cloning Output Paper Phase Physiology Production Productivity Proteins RNA Rana Reagent Reporting Research Research Personnel Resources Ruta System Systems Biology Terminator Codon Testing Time Tissues United States National Institutes of Health Voting Wood material Xenopus Xenopus laevis Yeasts cost epigenomics expression vector functional genomics genome-wide genomic tools in vivo innovation meetings nuclear transfer screening tool vector

项目摘要

We propose to generate, validate and distribute the second version of the Xenopus ORFeome, which we define as a fully sequenced, validated set of Xenopus cDNA clones containing one each of every open reading frame (ORF) encoded in the genome, in a format in which any ORF sequence can be easily transferred using recombineering into a diverse array of expression vectors. This one set of reagents will greatly decrease the time to characterize any protein in the myriad functional assays carried out by the entire Xenopus community. But most importantly, an ORFeome set will allow high-throughput in vivo functional-genomic screening in manner currently not feasible, which is a particular strength of the Xenopus system where functional screens have been the basis of a number of fundamental biological observations. Our consortium completed the first phase of this project where we moved 90% of the available full-length cDNA clones to Gateway compatible vectors. However, because of limitations of the original Xenopus EST projects this represents only half of all of the ORFs in the genome. In this project we will de novo clone the remaining Xenopus ORFs to a Gateway donor vector to generate the only available cDNA clones for over half the genome. These clones will be made available, without restriction, to researchers worldwide. The Xenopus community is fully supportive of this project and at a recent PI meeting (MBL (Woodshole), September 2015) the ORFeome was voted as the Top priority of needed Xenopus resources.

我们建议生成，验证和分发Xenopus的第二版 Orfeome，我们将其定义为经过完全测序的，经过验证的Xenopus cDNA克隆在基因组中编码的每个开放式阅读框（ORF）中包含一个使用重新组合可以轻松传递任何ORF序列的格式各种表达向量。这套试剂将大大降低是时候表征整个全功能测定中的任何蛋白质 Xenopus社区。但最重要的是，Orfeome套件将允许高通量当前不可行的方式，体内功能基因组筛选，这是一个特定的功能屏幕一直是A的Xenopus系统的强度基本生物学观察的数量。我们的财团完成了第一阶段在这个项目中，我们将90％的全长cDNA克隆移至Gateway 兼容的向量。但是，由于原始Xenopus EST项目的局限性这仅代表基因组中所有ORF的一半。在这个项目中，我们将从头开始克隆其余的xenopus orfs到网关供体向量，以生成唯一的可用的cDNA克隆，用于一半以上的基因组。这些克隆将被提供，不受限制地对全球研究人员进行限制。 Xenopus社区完全支持在该项目以及最近一次的PI会议（MBL（Woodshole），2015年9月） Orfeome被选为所需Xenopus资源的首要任务。