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.
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