Beyond GFP and aequorin: Ocean-wide study of fluorescent and luminous proteins

超越 GFP 和水母发光蛋白：荧光和发光蛋白的全海洋研究

• 批准号: 7133657
• 负责人: MIKHAIL V MATZ
• 金额: $ 27.62万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-09-01 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7133657
• 关键词: Xenopus; animal extract; aquatic organism; cell line; chemical stability; embryo /fetus; expression cloning; fluorescence; fluorescent dye /probe; genetic library; genetic screening; luminescence; mass screening; microorganism culture; protein purification; protein quantitation /detection; protein structure function; proteomics; transfection /expression vector

DESCRIPTION (provided by applicant): In vivo labeling and detection technologies based on genetically encoded fluorescent and luminescent markers is indispensable for modem biomedical science. We will clone proteins responsible for fluorescence and luminescence from a wide variety of organisms found in the open ocean and deep sea, and assess the usefulness of these proteins for biotechnology applications by expressing them in bacteria, mammalian cell culture and Xenopus laevis embryos. We are well poised to carry out such a project. Both our laboratories have regular access to oceanic and deep-sea animals through participation in research cruises sponsored by other agencies, including expeditions specifically equipped for studying physiology and ecology of oceanic fluorescence and luminescence. Our sampling methods include plankton towing, blue water SCUBA diving, midwater trawling and submersible/ROV operations. A collection of potential cloning targets has been already assembled that includes spectacular taxonomic diversity of animals, ranging from radiolarians and jellyfish to sea urchins and fish. This collection will be further augmented during upcoming research cruises in Caribbean, Eastern and Western Pacific. In the same time, we possess extensive experience in cloning, expression and analysis of fluorescent (Matz) and luminescent (Haddock) proteins. The guaranteed result will be identification of a series of novel calcium-dependent photoproteins, as well as GFP-like proteins possessing combinations of biophysical, biochemical and optical features that are not found in thoroughly sampled Anthozoa (such as far-red fluorescence, true monomeric state and rapid maturation). Our ultimate goal, however, is to identify completely novel (i.e., sharing no homology with the previously cloned ones) genetically encoded fluorescent and luminescent markers which may become a basis for the new generation of in vivo labeling and detection technologies.

描述（由申请人提供）：基于遗传编码的荧光和发光标记的体内标记和检测技术对于现代生物医学科学来说是必不可少的。我们将克隆蛋白质蛋白质，从而从开阔的海洋和深海中发现的各种生物产生荧光和发光，并通过在细菌，哺乳动物细胞培养和xenopus laevis embryos中表达这些蛋白质对生物技术应用的有用性。我们准备进行这样一个项目。我们的两个实验室都可以通过参与其他机构赞助的研究巡游定期进入海洋和深海动物，包括专门研究海洋荧光和发光生理学和生态学的探险。我们的抽样方法包括浮游生物牵引，蓝色水肺潜水，中间拖网和潜水/ROV操作。已经组装了一系列潜在的克隆靶标，其中包括动物的壮观分类多样性，从放射性鱼类和水母到海胆和鱼类。在即将到来的加勒比海，东太平洋和西太平洋的研究巡航期间，该系列将进一步增加。同时，我们在克隆，表达和分析荧光（MATZ）和发光（HADDOCK）蛋白方面具有丰富的经验。保证的结果将是鉴定一系列新型钙依赖性的光蛋白以及具有生物物理，生化和光学特征组合的GFP样蛋白，这些蛋白在彻底取样的Anthozoa中找不到（例如远红色的荧光，真实的单元状态和快速成熟）。然而，我们的最终目标是识别完全新颖的（即与先前克隆的荧光灯共享同源性）遗传编码的荧光和发光标记物，这可能成为新一代的体内标记和检测技术的基础。