Functional Analysis of Filamentous Fungi

丝状真菌的功能分析

• 批准号: 6958171
• 负责人: Jay C. Dunlap
• 金额: $ 114.98万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-04-01 至 2009-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6958171
• 关键词: Neurospora; functional /structural genomics; fungal genetics; fungal proteins; gene mutation; microorganism resource /registry; phenotype; polymerase chain reaction; protein structure function

The overall goal of the four interdependent projects in this Program Project is to carry out functional genomics, annotation, and expression analyses of Neurospora crassa, the filamentous fungus that has become a model for the assemblage of over 250,000 species of non-yeast fungi. Most Neurospora genes have no homologs in yeasts and nearly 40% have no strong homologs in any organism to date, suggesting that examination of the functions of these genes will both novel and informative. Neurospora is an important model for basic research in eukaryotes, and fungi allied to Neurospora include significant animal and plant pathogens and industrial strains yielding antibiotics, chemicals, enzymes, and pharmaceuticals. This subproject will pursue the systematic disruption of genes through targeted gene replacements, preliminary phenotyping of these strains, and their distribution to the scientific community at large. The 43 Mb Neurospora genome is completely sequenced, and automated annotation using programs trained on Neurospora genes predict 10,082 proteins. Phenotypes are now associated with about 10% of these, and roughly half of the genes have no strong sequence homologs in other organisms; thus, there are no clues to their function(s). Furthermore, it is likely that functions of some genes sharing sequence similarity with genes in other organisms will be modified slightly or greatly, reflecting the novel biology of filamentous fungi. Our long term goal is to create gene knockouts in all of these genes as a first step to determining each gene's function(s), and the immediate goal of the five Specific Aims of Project #1 is to facilitate this. In Specific Aim #1, we will use PCR, homologous recombination in yeast, and long range PCR to generate constructions (knockout cassettes) that can be used to knockout all of the genes in Neurospora. In Specific Aim #2, we will carry out gene replacements in Neurospora to generate unambiguous functional knockouts. In Specific Aim #3, we will carry out a basic phenotypic characterization of all the mutants. In Specific Aim #4, we will deposit the novel strains in the Fungal Genetics Stock Center which will maintain the stocks and distribute them to the scientific community at large. This effort will help to anchor genomic exploration in the Kingdom of the Fungi.

该计划项目中四个相互依存项目的总体目标是进行功能性基因组学，注释和表达分析，对神经孢子虫（Neurospora Crassa）进行，这是一种丝状真菌，已成为250,000种超过250,000种非偶然真菌的模型。大多数神经孢子基因在酵母中没有同源物，迄今为止，近40％的人在任何生物体中都没有强大的同源物，这表明研究这些基因的功能将既新颖又有信息。 Neurospora是真核生物基础研究的重要模型，与Neurospora结盟的真菌包括重要的动物和植物病原体以及产生抗生素，化学物质，酶和药物的工业菌株。 该副本将通过靶向基因的替代品，这些菌株的初步表型以及它们分布到整个科学群落来追求基因的系统破坏。 43 MB神经孢子基因组已完全测序，并使用接受神经孢子基因训练的程序预测10,082种蛋白质自动注释。现在，表型与其中约10％有关，大约一半的基因在其他生物体中没有强序同源物。因此，它们的功能没有线索。此外，某些基因与基因共享序列相似性的功能很可能 在其他生物中，将稍作或大量修饰，反映出丝状真菌的新生物学。我们的长期目标是在所有这些基因中创建基因敲除作为确定每个基因功能的第一步，而项目＃1的五个特定目标的直接目标是促进这一点。在特定的目标＃1中，我们将使用PCR，酵母中的同源重组，以及远距离PCR来生成构造（基因敲除盒），可用于敲除神经孢子中的所有基因。在特定的目标＃2中，我们将在Neurospora中进行基因替代，以产生明确的功能敲除。在特定的目标＃3中，我们将对所有突变体进行基本的表型表征。在特定的目标＃4中，我们将将新颖菌株存放在真菌遗传库存中心中，该菌株将维护股票并分发 他们到整个科学界。这项工作将有助于在真菌王国中锚定基因组探索。