Focus: The interface between data collection and data processing in cryo-EM.

We present a new software package called Focus that interfaces cryo-transmission electron microscopy (cryo-EM) data collection with computer image processing. Focus creates a user-friendly environment to import and manage data recorded by direct electron detectors and perform elemental image processing tasks in a high-throughput manner while new data is being acquired at the microscope. It provides the functionality required to remotely monitor the progress of data collection and data processing, which is essential now that automation in cryo-EM allows a steady flow of images of single particles, two-dimensional crystals, or electron tomography data to be recorded in overnight sessions. The rapid detection of any errors that may occur greatly increases the productivity of recording sessions at the electron microscope.

[1]  Anchi Cheng,et al.  Automated molecular microscopy: the new Leginon system. , 2005, Journal of structural biology.

[2]  Sebastian Scherer,et al.  2dx_automator: implementation of a semiautomatic high-throughput high-resolution cryo-electron crystallography pipeline. , 2014, Journal of structural biology.

[3]  David J. Fleet,et al.  cryoSPARC: algorithms for rapid unsupervised cryo-EM structure determination , 2017, Nature Methods.

[4]  Wen Jiang,et al.  Antibody-Based Affinity Cryo-Electron Microscopy at 2.6 Å Resolution , 2016, bioRxiv.

[5]  David A Agard,et al.  Asynchronous data acquisition and on-the-fly analysis of dose fractionated cryoEM images by UCSFImage. , 2015, Journal of structural biology.

[6]  D. Agard,et al.  Electron counting and beam-induced motion correction enable near atomic resolution single particle cryoEM , 2013, Nature Methods.

[7]  K. Grünewald,et al.  Electron Cryotomography of Tula Hantavirus Suggests a Unique Assembly Paradigm for Enveloped Viruses , 2010, Journal of Virology.

[8]  Wen Jiang,et al.  EMAN2: an extensible image processing suite for electron microscopy. , 2007, Journal of structural biology.

[9]  Dmitry Lyumkis,et al.  Likelihood-based classification of cryo-EM images using FREALIGN. , 2013, Journal of structural biology.

[10]  John W Sedat,et al.  UCSF tomography: an integrated software suite for real-time electron microscopic tomographic data collection, alignment, and reconstruction. , 2007, Journal of structural biology.

[11]  Nikolaus Grigorieff,et al.  FREALIGN: high-resolution refinement of single particle structures. , 2007, Journal of structural biology.

[12]  Christopher Irving,et al.  Appion: an integrated, database-driven pipeline to facilitate EM image processing. , 2009, Journal of structural biology.

[13]  Rafael Fernandez-Leiro,et al.  A pipeline approach to single-particle processing in RELION , 2016, bioRxiv.

[14]  Marcus A. Brubaker,et al.  Alignment of cryo-EM movies of individual particles by optimization of image translations. , 2014, Journal of structural biology.

[15]  D. Agard,et al.  MotionCor2: anisotropic correction of beam-induced motion for improved cryo-electron microscopy , 2017, Nature Methods.

[16]  Michael Schatz,et al.  Four-Dimensional Cryo Electron Microscopy at Quasi Atomic Resolution: "IMAGIC 4D" , 2012 .

[17]  Xiangyan Zeng,et al.  2dx_merge: data management and merging for 2D crystal images. , 2007, Journal of structural biology.

[18]  J. McIntosh,et al.  The Molecular Architecture of Axonemes Revealed by Cryoelectron Tomography , 2006, Science.

[19]  N Grigorieff,et al.  Frealign: An Exploratory Tool for Single-Particle Cryo-EM. , 2016, Methods in enzymology.

[20]  Edward H Egelman,et al.  The iterative helical real space reconstruction method: surmounting the problems posed by real polymers. , 2007, Journal of structural biology.

[21]  C O S Sorzano,et al.  Scipion: A software framework toward integration, reproducibility and validation in 3D electron microscopy. , 2016, Journal of structural biology.

[22]  Sjors H.W. Scheres,et al.  RELION: Implementation of a Bayesian approach to cryo-EM structure determination , 2012, Journal of structural biology.

[23]  W. Kühlbrandt.  The Resolution Revolution , 2014, Science.

[24]  F. Sigworth.  A maximum-likelihood approach to single-particle image refinement. , 1998, Journal of structural biology.

[25]  J R Kremer,et al.  Computer visualization of three-dimensional image data using IMOD. , 1996, Journal of structural biology.

[26]  Kai Zhang,et al.  Gctf: Real-time CTF determination and correction , 2015, bioRxiv.

[27]  Daniel Castaño-Díez,et al.  Dynamo: a flexible, user-friendly development tool for subtomogram averaging of cryo-EM data in high-performance computing environments. , 2012, Journal of structural biology.

[28]  Roberto Marabini,et al.  MRC2014: Extensions to the MRC format header for electron cryo-microscopy and tomography , 2015, Journal of structural biology.

[29]  S. Harrison,et al.  Lipid–protein interactions in double-layered two-dimensional AQP0 crystals , 2005, Nature.

[30]  Irina Gutsche,et al.  SPRING - an image processing package for single-particle based helical reconstruction from electron cryomicrographs. , 2014, Journal of structural biology.

[31]  N. Grigorieff,et al.  Quantitative characterization of electron detectors for transmission electron microscopy. , 2013, Journal of structural biology.

[32]  S Rawson,et al.  Methods to account for movement and flexibility in cryo-EM data processing , 2016, Methods.

[33]  J. Frank,et al.  Structure and assembly model for the Trypanosoma cruzi 60S ribosomal subunit , 2016, Proceedings of the National Academy of Sciences.

[34]  S J Ludtke,et al.  Single-Particle Refinement and Variability Analysis in EMAN2.1. , 2016, Methods in enzymology.

[35]  Friedrich Förster,et al.  TOM software toolbox: acquisition and analysis for electron tomography. , 2005, Journal of structural biology.

[36]  Mindy I. Davis,et al.  Breaking Cryo-EM Resolution Barriers to Facilitate Drug Discovery , 2016, Cell.

[37]  J. Kowal,et al.  Robust image alignment for cryogenic transmission electron microscopy. , 2017, Journal of structural biology.

[38]  J. Briggs,et al.  Implementation of a cryo-electron tomography tilt-scheme optimized for high resolution subtomogram averaging , 2017, Journal of structural biology.

[39]  Xiangyan Zeng,et al.  2dx--user-friendly image processing for 2D crystals. , 2007, Journal of structural biology.

[40]  S. Stagg,et al.  Automated batch fiducial-less tilt-series alignment in Appion using Protomo. , 2015, Journal of structural biology.

[41]  N. Grigorieff,et al.  CTFFIND4: Fast and accurate defocus estimation from electron micrographs , 2015, bioRxiv.

[42]  Nikolaus Grigorieff,et al.  Measuring the optimal exposure for single particle cryo-EM using a 2.6 Å reconstruction of rotavirus VP6 , 2015, eLife.

[43]  David N Mastronarde,et al.  Automated electron microscope tomography using robust prediction of specimen movements. , 2005, Journal of structural biology.