THREE DIMENSIONAL RECONSTRUCTIONS OF ARCHAEA

古细菌的三维重建

• 批准号: 8170822
• 负责人: NORMAN R PACE
• 金额: $ 3.74万
• 依托单位: UNIVERSITY OF COLORADO
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2011-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8170822
• 关键词: Archaea; Bacteria; Biochemical; Bioinformatics; Biological; Biological Preservation; Cell membrane; Cell model; Cells; Complement; Computer Retrieval of Information on Scientific Projects Database; Computer Simulation; Computer software; Culture Media; Custom; DNA; DNA Probes; Eukaryota; Exposure to; Freeze Substitution; Freezing; Funding; Genetic Transcription; Grant; Heat-Shock Response; Heavy Metals; Hydrogen Peroxide; Image; Institution; Knowledge; Metabolic; Methods; Molecular; Morphology; Organism; Oxidative Stress; Phylogenetic Analysis; Preparation; Process; Prokaryotic Cells; Protocols documentation; Research; Research Personnel; Resources; Ribosomes; Salmonella; Sampling; Source; Staining method; Stains; Structure; Sulfolobus; Sulfolobus solfataricus; Tomogram; Translations; Trees; United States National Institutes of Health; Work; cellular imaging; instrumentation; pressure; rapid technique; reconstruction; sugar; 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. Work conducted in previous years has resulted in sample preparation protocols that routinely yield very high quality full-cell tomograms of these two organisms. Custom software created by Dr. Robertson automatically segments the tomographic image stacks for the major biomolecules which stain with heavy metals, (ribosomes, cell membrane, etc) in Salmonella and Sulfolobus. The automatic segmentation software provides the means to enumerate these biomolecules as the organisms respond to a variety of metabolic states such as heat shock, oxidative stress (exposure to hydrogen peroxide), or changes in growth medium (eg sugar/no sugar). The auto-segmentation software also generates structural bioinformatic models of the cells which may be used for in silico experimentation. The tomographic work has been complemented with immuno-EM of Lowicryl embedded Sulfolobus, has been successfully probed for DNA.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这不一定是调查员的机构。 所有已知生物的系统发育树都包括三个不同的分支：欧洲，细菌和古细菌。 古细菌分支最近才被公认为是一个连贯的组，在系统发育上与细菌不同。 我们对原核细胞（古细菌和细菌）的精细结构的了解明显少于我们对它们的生化和分子组织的了解。 这主要是由于保存方法的局限性，并且在某种程度上是仪器的限制。 快速冷冻和冻结固定的现代技术的进步已经克服了其中的一些问题，从而优雅地保存了一些原核生物。 由于分子研究表明，在转录，翻译和其他与DNA相关的处理机制方面，古细菌与真核生物相比与细菌有更多的共同点，因此这些生物的形态可能会产生大量的生物学信息。 高压冷冻，冻结取代的Sulfolobus solfataricus的全细胞图像已通过串行部分断层扫描捕获。 作为对照，沙门氏菌细菌的全细胞图像已经以相同的方式制备。 往年进行的工作导致样品制备方案通常会产生这两个生物的非常高质量的全细胞中层图。由Robertson博士创建的自定义软件自动将层析成分图像堆栈列为主要生物分子，这些生物分子用沙门氏菌和硫bus中的重金属（核糖体，细胞膜等）染色。 自动分割软件提供了列举这些生物分子的手段，因为生物体对各种代谢状态（例如热休克，氧化应激（暴露于过氧化氢）或生长培养基的变化（例如糖/NO糖）的变化（例如，糖/NO糖）。 自动分割软件还会生成细胞的结构生物信息学模型，这些模型可用于计算机实验中。 层析成像工作已与Lowicryl嵌入的磺胺的免疫EM相辅相成，已成功探测DNA。