The Brazil School for Single Particle Cryo-EM is an intensive hands-on course/workshop, in which theory and practice are tightly intertwined. The spectrum of topics covered suits attendees that are new to the field as well as those for whom the technique is already of growing importance in their research and who wish to acquire an in-depth understanding of the methodology. Topics range from data collection and three-dimensional reconstruction techniques to the final interpretation of the cryo-EM maps by (flexible) fitting of X-ray structures. The last days of the meeting will be devoted to advanced topics such as dealing with heterogeneous data sets ("4D" cryo electron microscopy) and high-resolution refinements.
Main Lecture Topics
• The Fourier transform in 1D - 2D - 3D
• Properties of the Fourier transform
• Specimen preparation for single-particle cryo-EM
• (Automatic) Particle selection
• Optimizing the EM data collection
• CTF correction
• Filtering and data normalization
• Alignment using correlation functions
• Multi-reference alignments
• Multivariate statistics and classification
• Angular reconstitution
• Three-dimensional reconstruction
• Projection matching
• Alignment refinements
• 3D refinements
• 3D resolution criteria
• 3D fitting of known X-ray structures (Chimera, Coot)
• Refining towards different 3D structures simultaneously ("4D" analysis)
• Using image processing software (IMAGIC, Frealign)
• Parallel processing
The attendees are expected to bring along a modern notebook computer onto which all necessary software will be installed. Most processing will take place in the context of the IMAGIC software system but other software packages such as Frealign will also be used. Cryo-EM data sets will be distributed for processing during the course. The participants may bring their own data sets for processing and discussion.
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