THREE DIMENSIONAL RECONSTRUCTIONS OF ARCHAEA

古细菌的三维重建

• 批准号: 7722829
• 负责人: NORMAN R PACE
• 金额: $ 2.76万
• 依托单位: UNIVERSITY OF COLORADO
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-08-01 至 2009-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7722829
• 关键词: Archaea; Bacteria; Biochemical; Biological; Biological Preservation; Cells; Computer Retrieval of Information on Scientific Projects Database; Culture Media; DNA; DNA Probes; Eukaryota; Eukaryotic Cell; Exclusion; Ficoll; Freeze Substitution; Freezing; Funding; Genetic Transcription; Glucose; Grant; Institution; Knowledge; Methods; Modeling; Molecular; Morphology; Nature; Nuclear; Organism; Phylogenetic Analysis; Polymers; Process; Prokaryotic Cells; Rate; Research; Research Personnel; Resources; Ribosomes; Salmonella; Source; Structure; Sucrose; Sulfolobus; Sulfolobus solfataricus; Tomogram; Translations; Trees; United States National Institutes of Health; Work; cellular imaging; instrumentation; pressure; rapid technique; reconstruction; tomography

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The phylogenetic tree of all known organisms comprises three distinct branches: Eucarya, Bacteria and Archaea. The Archaeal branch was only recently recognized as a coherent group that is phylogenetically distinct from Bacteria. Our knowledge of the fine structure of prokaryotic cells (Archaea and Bacteria) is significantly less than what we know about their biochemical and molecular organization. This is due primarily to limitations in preservation methods and, to some extent, in instrumentation. Advances in modern techniques of rapid freezing and freeze-substitution have overcome some of these problems, yielding elegant preservation of some prokaryotic organisms. Since molecular studies suggest that Archaea have more in common with Eukaryotes than with Bacteria, in terms of transcription, translation and other DNA related processing mechanisms, the morphology of these organisms may yield a wealth of biological information. Whole cell images of high-pressure frozen, freeze-substituted Sulfolobus solfataricus have been captured via serial section tomography. As a control, whole cell images of the bacterium Salmonella have been prepared in an identical manner. The whole cell tomograms have allowed the enumeration of several different structures in Sulfolobus, at least one of which varies according to the presence or absence of glucose in the medium in which the cells are grown. DNA preservation, previously a problem, is currently quite good with the addition of the sucrose polymer Ficoll-70 to the growth medium just prior to freezing. Lowicryl embedded Sulfolobus has been successfully probed for DNA and verified by tomographic ribosome exclusion models. Work will continue on the characterization of nuclear body structure of Sulfolobus and Salmonella during the next year. The principal rate determining step going forward is the labor intensive nature of segmentation.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目和 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 所有已知生物体的系统发育树包括三个不同的分支：真核生物、细菌和古细菌。 古细菌分支直到最近才被认为是一个在系统发育上与细菌不同的连贯群体。 我们对原核细胞（古细菌和细菌）精细结构的了解远远少于我们对其生化和分子组织的了解。 这主要是由于保存方法的限制以及在某种程度上仪器的限制。 现代快速冷冻和冷冻替代技术的进步已经克服了其中一些问题，从而对一些原核生物进行了优雅的保存。 由于分子研究表明古细菌与真核生物比细菌有更多的共同点，在转录、翻译和其他 DNA 相关加工机制方面，这些生物体的形态可能会产生丰富的生物信息。 高压冷冻、冷冻替代的硫磺硫化叶菌的全细胞图像已通过连续切片断层扫描捕获。 作为对照，以相同的方式制备了沙门氏菌的全细胞图像。 全细胞断层扫描可以枚举硫化叶菌中的几种不同结构，其中至少一种结构根据细胞生长培养基中是否存在葡萄糖而变化。 DNA 保存以前是一个问题，但目前通过在冷冻前向生长培养基中添加蔗糖聚合物 Ficoll-70 可以得到很好的保存。 Lowicryl 嵌入的硫化叶菌已成功探测 DNA 并通过断层扫描核糖体排除模型进行验证。 明年将继续研究硫化叶菌和沙门氏菌核体结构的特征。 未来决定利率的主要步骤是细分的劳动密集型性质。