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. Structural visualization using electron microscopy has provided many insights into the composition, arrangement and mechanism of macromolecular assemblies. By preserving samples in stain or vitreous ice it is possible to image them as discrete particles, and from these images generate three-dimensional structures. This 'single-particle'approach suffers from two major shortcomings;it requires an initial model to reconstitute 2D data into a 3D volume, and it often fails when faced with conformational variability. Random conical tilt (RCT) and orthogonal tilt (OTR) are methods developed to overcome these problems, but the data collection required, particularly in ice, is difficult and tedious.
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