DETERMINATION OF CHROMOSOME STRUCTURE FROM EM TOMOGRAPHY

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。Sedat实验室开发的电子显微镜（EM）层析成像技术提供了最小扰动固定和染色（后期）染色体的高分辨率数据。硬件的理论分辨率应该足以使染色体的错综复杂的结构可视化，它被认为是一个由单链DNA组成的紧密交织的物体。然而，典型的三维层析重建具有明显的云状结构，这使得单链DNA的识别非常困难。虽然一些分辨率的损失是固有的断层扫描重建，大多数的退化是由于背景染色和非特异性染色的DNA，这是保留在断层扫描重建。我们利用来自RBVI的Chimera建模软件来可视化一些处理过的EM体积数据，并与RBVI的Tom Goddard合作，改进了Chimera的“体积查看器”扩展功能，以便更好地可视化我们的EM数据。今年，我们一直在使用我们开发的基于3D小波的滤波器（临时专利文件为10/03）来研究染色体结构。我们最近向J. Microsc提交了一篇论文“一种新的基于三维小波的滤波器，用于在嘈杂的生物数据中显示特征”，作者是W. C. Moss， S. Haase， J. M. Lyle， D. a . Agard， J. W. Sedat。

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. Electron Microscope (EM) tomographic techniques developed in the Sedat lab provide high resolution data of minimally perturbed fixed and stained (anaphase) chromosomes. The theoretical resolution of the hardware should be sufficient to visualize the convoluted structure of the chromosome, which is expected to appear as a tightly interwoven object composed of a single strand of DNA. However, the typical three-dimensional tomographic reconstruction has an apparent cloud-like structure, which makes identification of a single strand of DNA very difficult. Although some loss of resolution is inherent in the tomographic reconstruction, most of the degradation is due to background staining and nonspecific staining of the DNA, which is retained in the tomographic reconstruction. We have utilized the Chimera modeling software from the RBVI to visualize some of the processed EM volume data, and have worked with Tom Goddard at the RBVI to improve the capabilities of the "Volume Viewer" extension to Chimera in order to better visualize our EM data. This year we have been using a 3D wavelet-based filter (provisional patent filed 10/03) that we have developed to study chromosome structure. We recently submitted a paper "A novel 3D wavelet based filter for visualizing features in noisy biological data", by W. C. Moss, S. Haase, J. M. Lyle, D. A. Agard, J. W. Sedat, to J. Microsc.