Regulatory Genomics: BAC-GFP Library of Control Genes

调控基因组学：BAC-GFP 对照基因文库

• 批准号: 7390337
• 负责人: ERIC H DAVIDSON
• 金额: $ 26.26万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-04-01 至 2010-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7390337
• 关键词: Appendix; Code; Communities; Complementary DNA; Data; Development; Embryo; Exons; Facility Construction Funding Category; Gene Expression; Gene Transfer; Genes; Genome; Genomics; Green Fluorescent Proteins; Internet; Laboratories; Larva; Letters; Libraries; Maps; Methods; Molecular Profiling; Pattern; Production; Proteins; Protocols documentation; Recombinants; Regulator Genes; Reporter; Research Personnel; Resources; Scientist; Sea Urchins; Site; Standards of Weights and Measures; Strongylocentrotus purpuratus; System; Techniques; Testing; Transcription factor genes; cDNA Library; egg; expression vector; genetic regulatory protein; homologous recombination; interest; programs; tool; transcription factor

We propose to develop a communal resource of BAG GFP expression vectors of all transcription factors expressed significantlyin the embryo of the sea urchin, Strongylocentrotus purpuratus, the genomic sequence of which will go on line shortly. Critically, as shown by our laboratory, BAG GFP recombinants can be injected into eggs and with high efficiency, and they express accurately, capturing the entire cis-regulatory systems intact. This greatly accelerates detailed cis-regulatory analyses. These recombinants can also be used to introduce exogenous regulatory gene products in temporally and spatiallyrestricted patterns within the embryo and larva. Both of these applications will greatly enhance analysis of how the genomic control systems which direct expression of regulatory genes prescribe early development. Specifically, we propose to isolate BAG recombinants for all relevant transcription factors (-100-200) in which the gene of interest is centrally located within the clone, and then to use standard homologous recombination techniques to replace the first exon of the endogenous gene with the coding sequence of the GFP reporter. An ongoing project within our laboratory has already established the expression profiles of most of the pertinent genes. We will then employ our methods for high throughput production of BAC-GFP recombinants in exonl of each gene, and build and authenticate each construct. GFP expression directed by each BAG recombinant will be analyzed to confirm faithful recapitulation of endogenous gene expression. We will also make BAG recombinants for any other gene encoding any kind of protein, that scientists in the community may request. All information needed by other investigators to facilitate access and promote the use of these tools will be made available on the Sea Urchin Genome web site that is currently administrated by our laboratory. Numerous letters from outside investigators attesting interest in this facility are included as an Appendix to this application.

我们建议开发一个所有转录因子的BAG GFP表达载体的公共资源