C.O.S. Sorzano, J. M. de la Rosa Trevín, J. Otón, J. J. Vega, J. Cuenca, A. Zaldívar-Peraza, J. Gómez-Blanco, J. Vargas, A. Quintana, R. Marabini, J. M. Carazo. Semiautomatic, high-throughput, high-resolution protocol for three-dimensional reconstruction of Single Particles in Electron Microscopy. Nanoimaging: Methods and Protocols. Methods in Molecular Biology, 950: 171-193. Eds. Alioscka Sousa, Michael Kruhlak. Humana Press. (2012)
In this chapter we describe the steps needed for reconstructing the three-dimensional structure of a macromolecular complex starting from its projections collected in electron micrographs. The concepts are shown through the use of Xmipp 3.0, a software suite specifically designed for the image processing of biological structures imaged with electron or X-ray microscopy. We illustrate the image processing workflow by applying it to the images of Bovine Papilloma virus published in Wolf et al. (Proc Natl Acad Sci USA 107:6298–6303, 2010). We show that in the case of high-quality, homogeneous datasets with a priori knowledge about the initial volume, we can have a high-resolution 3D reconstruction in less than 1 day using a computer cluster with only 32 processors.
Single Particle Analysis, High-throughput, Xmipp