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Semiautomatic, High-Throughput, High-Resolution Protocol for Three-Dimensional Reconstruction of Single Particles in Electron Microscopy

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Nanoimaging

Part of the book series: Methods in Molecular Biology ((MIMB,volume 950))

Abstract

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.

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Acknowledgements

C.O.S. Sorzano is a recipient of a Ramón y Cajal fellowship awarded by the Spanish Ministry of Science and innovation (Micinn). J. Vargas and J. Otón are Juan de la Cierva fellows (Micinn). This work has been partly funded by grants CSD2006-0002 (Spanish Ministry of Education), ACI2009-10220 (Micinn), BIO2010-16566 (Micinn), and PIF08-020-2 (CSIC).

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Authors and Affiliations

  1. Biocomputing Unit, National Center of Biotechnology (CSIC), Universidad Autónoma de Madrid, Madrid, Spain

    Carlos Oscar Sorzano, J. M. de la Rosa Trevín, J. Otón, J. J. Vega, J. Cuenca, A. Zaldívar-Peraza, J. Gómez-Blanco, J. Vargas, A. Quintana & José María Carazo

  2. Escuela Politecnica Superior, Universidad Autónoma de Madrid, Madrid, Spain

    Roberto Marabini

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  1. Carlos Oscar Sorzano
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  2. J. M. de la Rosa Trevín
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  3. J. Otón
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  4. J. J. Vega
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  5. J. Cuenca
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  6. A. Zaldívar-Peraza
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  7. J. Gómez-Blanco
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  8. J. Vargas
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  9. A. Quintana
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  10. Roberto Marabini
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  11. José María Carazo
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Corresponding author

Correspondence to Carlos Oscar Sorzano .

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Editors and Affiliations

  1. Nat. Inst. Biomedical Imaging & Bioengin, National Institutes of Health, South Drive 13, Bethesda, 20892, Maryland, USA

    Alioscka A. Sousa

  2. , National Cancer Institute, National Institutes of Health, Center Drive 10, Bethesda, 20892, USA

    Michael J. Kruhlak

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Sorzano, C.O. et al. (2013). Semiautomatic, High-Throughput, High-Resolution Protocol for Three-Dimensional Reconstruction of Single Particles in Electron Microscopy. In: Sousa, A., Kruhlak, M. (eds) Nanoimaging. Methods in Molecular Biology, vol 950. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-137-0_11

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  • DOI: https://doi.org/10.1007/978-1-62703-137-0_11

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-136-3

  • Online ISBN: 978-1-62703-137-0

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