Comprehensive Resource for the Drosophila 4th chromosome

果蝇第四染色体综合资源

基本信息

批准号：
10625841
负责人：
STUART J NEWFELD
金额：
$ 68.22万
依托单位：
ARIZONA STATE UNIVERSITY-TEMPE CAMPUS
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-06-01 至 2025-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10625841
关键词：
ANK2 gene Adult Advisory Committees Age related macular degeneration Ankyrins BCL9 gene Basic Science Behavior Biological Biological Assay Biology Brain Categories Central Nervous System Chromosomes Clone Cells Clustered Regularly Interspaced Short Palindromic Repeats Code Collaborations Collection Communities Complementary DNA Defect Development Disease Drosophila genus Drosophila melanogaster Erinaceidae Eye FOXL1 gene Family Funding GDF8 gene Gene Expression Regulation Genes Genetic Recombination Genome Germ Lines Goals Health Heart Homologous Gene Human Letters Long QT Syndrome Longevity Methods Modeling Molecular Molecular Analysis Molecular Genetics Muscle Mutation Mutation Analysis Neuronal Differentiation Neurons Pattern Proteins RNA Reporter Genes Research Research Personnel Resource Development Resources Scientist Signal Pathway System Technology Tissues Transforming Growth Factor beta United States National Institutes of Health Untranslated RNA Work arm cooking fly frontier gain of function genetic analysis inhibin innovation interest loss of function loss of function mutation nervous system disorder neurotransmission synergism tool translational applications web site

项目摘要

For 125 years, Drosophila melanogaster has led the way in the genetic analysis of biological questions. The 4th chromosome is the final frontier for genetic analysis in Drosophila. Small and devoid of recombination the 4th has long been ignored. Nevertheless it contains 105 genes. 74% of the protein coding genes have human homologs and 68% of these have a disease association. For example Eyeless belongs to the PAX/RAX family where somatic loss of human RAX2 leads to age-related macular degeneration. Mutations in human ANK2, homolog of Ankyrin are the primary cause of congenital Long QT syndrome. A complete understanding of health requires the examination of these genes. To advance this effort, the PI recently generated unique chromosomes for the study of marked single cell clones (MARCM) carrying mutations on the 4th. Here he proposes to collaborate with colleagues at IU and UMN to facilitate the genetic and molecular analyses of every gene on the 4th. The Specific Aim is to generate a comprehensive resource for the Drosophila 4th chromosome. The resource will contain roughly 730 stocks divided into five collections. 1. FRT with a CRISPR mutation for each of the 79 protein coding genes for loss of function studies and MARCM (two mutations per gene = 158 stocks) 2. FRT with a CRISPR mutation for each of 26 noncoding RNAs for loss of function and MARCM (26 stocks). 3. Conversion of protein coding genes and noncoding RNAs with an existing MIMIC to T2A.GAL4:GFP and insertion of a CRIMIC that has T2A.GAL4:GFP in the remainder for fluorescent tagging, as reporter genes (protein or RNA) and gain of function studies (120 stocks). 4. Gain of function stocks composed of: a) UASt and UASp/UASz for each protein coding gene and non-coding RNA and b) UASt and UASp/UASz for the two closest human cDNAs for conserved protein coding genes and noncoding RNAs (400 stocks). 5. Balancer chromosomes and auxiliary chromosomes for clonal analyses such as FRT-GAL80 for MARCM, FRT-attP for designer clones and FRT-ovoD for germ line clones with/without UAS.FLP (20 stocks). The resource will enable: loss and gain of function assays, tissue-specific and temporal gene regulation in somatic and germ line tissues, the tracking of tagged RNA and proteins and all manner of genetic analyses. To assist in prioritizing tasks, we have an Advisory Committee and will accept community input. Characterization of new stocks takes place within the molecular genetics expertise of the investigators. In addition to facilitating basic research on development and disease, our resource will have direct translational application to understanding human health. For example, T2A.GAL4:GFP insertions that disrupt the fly gene and result in the expression of GAL4 in native patterns can be combined with UAS human cDNA stocks for the analysis of “humanized” disease or treatment models. This resource will be made readily available to researchers to advance our understanding of biology and conserved molecular mechanisms underlying human health.

在125年的时间里，果蝇Melanogaster一直领导着生物学问题的遗传分析。第四染色体是果蝇遗传分析的最终前沿。小而没有重组第四长期以来一直被忽略。然而，它包含105个基因。 74％的蛋白质编码基因具有人类同源物和其中68％具有疾病关联。例如，无眼属于Pax/Rax家族人类RAX2的体细胞丧失导致与年龄相关的黄斑变性。人类ANK2中的突变， Ankyrin的同源物是先天性长QT综合征的主要原因。完全理解健康需要检查这些基因。为了促进这项工作，PI最近产生了独特的用于研究第四个突变的明显单细胞克隆（MARCM）的染色体。他在这里与IU和UMN的同事合作的建议，以促进遗传和分子分析第四个基因。具体的目的是为果蝇第四生成全面的资源染色体。该资源将包含大约730个股票，分为五个收藏。 1。FRT与CRISPR 79种蛋白质编码基因中的每一个突变，用于丧失功能研究和MARCM（每个突变每个突变基因= 158个股票）2。frt具有26个非编码RNA中的CRISPR突变，用于功能损失和 Marcm（26个股票）。 3。蛋白质编码基因和非编码RNA的转换具有现有模拟 t2a.gal4：GFP和犯罪插入t2a.gal4：gfp的案件，用于荧光标签，作为报告基因（蛋白质或RNA）和功能研究的增益（120个股票）。 4。功能库存的收益由：a）a）uast和uasp/uasz组成，用于每个蛋白质编码基因和非编码RNA以及b）uast和uast UASP/UASZ用于两个最接近的人cDNA，用于保守的蛋白质编码基因和非编码RNA（400 股票）。 5。用于克隆分析的均衡染色体和辅助染色体，例如frt-gal80 Marcm，用于设计师克隆的FRT-ATTP和带有UAS.FLP（20股）的生物线克隆的FRT-ovod。资源将实现：功能分析的损失和获取，组织特异性和临时基因调节体细胞和生殖线组织，标记的RNA和蛋白质的跟踪以及各种遗传分析。到协助优先考虑任务，我们有一个咨询委员会，并将接受社区意见。表征在研究人员的分子遗传学专业知识中进行了新股票。除了促进关于发展和疾病的基础研究，我们的资源将直接转化为了解人类健康。例如，t2a.gal4：破坏苍蝇基因并导致的GFP插入 GAL4在天然模式中的表达可以与UAS人类cDNA库存合并，以分析 “人性化”疾病或治疗模型。该资源将很容易为研究人员提供促进我们对生物学的理解，并构成人类健康的分子机制。