TranSPHIRE: automated and feedback-optimized on-the-fly processing for cryo-EM

Stabrin M, Schoenfeld F, Wagner T, Pospich S, Gatsogiannis C, Raunser S (2020) Nat Commun

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Single particle cryo-EM requires full automation to allow high-throughput structure determination. Although software packages exist where parts of the cryo-EM pipeline are automated, a complete solution that offers reliable on-the-fly processing, resulting in highresolution structures, does not exist. Here we present TranSPHIRE: A software package for fully-automated processing of cryo-EM datasets during data acquisition. TranSPHIRE transfers data from the microscope, automatically applies the common pre-processing steps, picks particles, performs 2D clustering, and 3D refinement parallel to image recording. Importantly, TranSPHIRE introduces a machine learning-based feedback loop to re-train its picking model to adapt to any given data set live during processing. This elegant approach enables TranSPHIRE to process data more effectively, producing high-quality particle stacks. TranSPHIRE collects and displays all metrics and microscope settings to allow users to quickly evaluate data during acquisition. TranSPHIRE can run on a single work station and also includes the automated processing of filaments.

TranSPHIRE pipeline: Upper register (solid line): Overview of the integrated TranSPHIRE pipeline and all automate processing steps. The pipeline includes file management tasks, i.e., parallelized data transfer, file compression, and file backup (gray); 2D processing, i.e., motion correction, CTF estimation, particle picking, 2D clustering, and 2D class selection (turquoise); and 3D processing, i.e., ab initio 3D reconstruction and 3D refinement (red). Additionally, the pipeline includes an automated feedback loop optimization to adapt picking to the current data set during runtime (purple). Lower register (dotted line): The SPHIRE software package forms the backend for TranSPHIRE and offers the tools used for 2D and 3D processing. SPHIRE includes additional tools for advanced processing, such as heterogeneity analysis and local resolution determination.
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