THREE DIMENSIONAL RECONSTRUCTIONS OF ARCHAEA

该子项目是利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得主要资金， 因此可以在其他CRISP条目中表示。所列机构为 中心，不一定是研究者的机构。 所有已知生物的系统发育树包括三个不同的分支：真核生物，细菌和细菌。 直到最近才认识到，肠分支是一个在遗传学上与细菌不同的连贯类群。 我们对原核细胞（细菌和细菌）的精细结构的了解远远少于我们对它们的生物化学和分子组织的了解。 这主要是由于保存方法的局限性，在某种程度上也是由于仪器的局限性。 现代快速冷冻和冷冻替代技术的进步已经克服了其中的一些问题，使一些原核生物得到了优雅的保存。 由于分子生物学研究表明，在转录、翻译和其他与DNA相关的加工机制方面，古细菌与真核生物比与细菌有更多的共同点，因此这些生物的形态学可能产生丰富的生物学信息。 全细胞图像的高压冷冻，冷冻替代硫磺硫化叶菌已被捕获通过连续切片断层扫描。 作为对照，以相同的方式制备细菌沙门氏菌的全细胞图像。 全细胞断层图像允许枚举硫化叶菌中的几种不同结构，其中至少一种根据细胞生长培养基中葡萄糖的存在或不存在而变化。 DNA保存，以前是一个问题，目前在冷冻前向生长培养基中加入蔗糖聚合物Ficoll-70是相当好的。 Lowicryl包埋的Sulfolobus已成功探测DNA，并通过层析核糖体排除模型进行了验证。 明年将继续对硫化叶菌和沙门氏菌的核体结构进行鉴定。 决定未来利率的主要步骤是分割的劳动密集性质。

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.