Cryptococcus genomic resources

侵袭性真菌感染的有效诊断和治疗是临床上尚未解决的主要挑战。带注释的 主要真菌病原体的基因组序列为医学真菌学的革命创造了机会 通过应用系统的方法。为了最大限度地发挥作用，基因组序列必须连接在一起 拥有全基因组的生物资源。全基因组敲除和标记基因集合的可用性 以及全球遗传相互作用图在促进该模型的研究进展方面起到了重要作用 酿酒酵母。病原真菌中相似资源的产生创造了 实验工具使全面的方法能够阐明真菌毒力的基础在 哺乳动物的宿主和开发药物。新生隐球菌是一种被包裹的芽生酵母，它能引起 真菌性脑膜炎，每年造成约200,000人死亡。新生隐孢子虫具有单倍体遗传学，同源 重组，以及优秀的疾病动物模型。在上一阶段，我们使用了生物转化和 长同源靶向构建以完成基因敲除收集，执行广泛的质量控制 工作，开发了在池中筛选文库的定量方法，并在 小鼠和体外实验。我们已经通过真菌基因向社区提供了不受限制的菌株 股票中心(FGSC)。这一资源已经在30多种出版物中使用。加速基因组 经过改进，我们建立了第一个针对新生葡萄球菌的短同源CRISPR-Cas9方法。我们还有 创建了一个有效的定位和纯化相结合的标签。初步工作表明，这些 可以大规模部署方法，以构建标记菌株集合。此外，我们还有 设计了一种新的方法，可以在新生葡萄球菌中实现CRISPR-Cas9联合筛查。充分利用这些优势 技术进步，我们将1)完成隐球菌全基因组标签的收集，包括生产 2)构建以毒力为中心的第一代遗传互作图谱 决定因素。实现第一个目标将使细胞生物学和/或生化研究成为可能 任何蛋白质。完成第二个目标将通过在簇中加入成员来使大部分基因去孤构化 具有已知功能的基因，同时定义新的模块/通路。这些信息将被整合到 可搜索并可通过网络访问的VEuPathDB数据库。这些新的基因组资源将共同加速 研究界理解和开发针对最常见的致病原因的治疗方法的能力 人类真菌性脑膜炎。

Effective diagnosis and treatment of invasive fungal infections is a major unmet clinical challenge. Annotated genomic sequences of the major fungal pathogens create an opportunity to revolutionize medical mycology through the application of systematic approaches. To be maximally useful, genome sequences must be coupled with genome-wide biological resources. The availability of genome-wide knockout and tagged gene collections as well as a global genetic interaction map have been instrumental in catalyzing research progress in the model yeast Saccharomyces cerevisiae. The generation of analogous resources in pathogenic fungi creates experimental tools enabling comprehensive approaches to elucidating the basis of fungal virulence in the mammalian host and to develop drugs. Cryptococcus neoformans is an encapsulated budding yeast that causes fungal meningitis, resulting in ~200,000 fatalities each year. C. neoformans has haploid genetics, homologous recombination, and excellent animal models of disease. In the last period, we used biolistic transformation and long homology targeting constructs to complete a gene knockout collection, performed extensive quality control work, developed quantitative methods for screening the library in pools, and performed pooled screens both in mice and in vitro. We have made the strains available without restriction to the community via the Fungal Genetic Stock Center (FGSC). This resource has already been used in over 30 publications. To accelerate genome modification, we developed the first short-homology CRISPR-Cas9 methods for C. neoformans. We have also created an effective combined localization and purification tag. Preliminary work demonstrates that these methods can be deployed at scale to enable construction of a tagged strain collection. In addition, we have devised a novel method that enables pooled CRISPR-Cas9 screens in C. neoformans. Capitalizing on these technical advances, we will 1) complete the Cryptococcus whole-genome tag collection including production of an image resource, and 2) construct a first-generation genetic interaction map focused on virulence determinants. Accomplishment of the first aim will enable the cell biological and/or biochemical investigations of any protein. Completion of the second aim will deorphanize large fraction genes through membership in clusters of genes with known functions while defining new modules/pathways. This information will be integrated into the searchable and web accessible VEuPathDB database. Together these new genomic resources will accelerate the ability of the research community to understand and develop therapies against the most common cause of human fungal meningitis.