subflow

Workflow Overview

↓ on-the-fly

Monitors incoming files for on-the-fly processing of movies, micrographs, and particles.
Aligns movie frames and estimates CTF values using Relion schemes (Burt et al., 2024).
Monitors preprocessed micrographs and picks filaments using crYOLO (Wagner et al., 2020).
Subtracts those filaments from micrographs using the lattice-subtraction scripts (Chai et al., 2022).
Filaments are split before subtraction to reduce artefacts induced by bending.
Allows for two rounds of filament picking and subtraction if requested.
Picks particles from subtracted micrographs using crYOLO (Wagner et al., 2019).
Allows viewing of micrographs and picked coordinates (if available) to monitor progress.

↑ on-the-fly

Prepares Relion .star files with the modified paths to the subtracted micrographs.
Extracts particles, imports them to cryoSPARC (Punjani et al., 2017).
Performs heterogeneous refinement in cryoSPARC with user-defined volumes to extract a good class.
Converts the particles back to Relion .star format using pyem (Asarnow et al., 2019).

Set up a fresh conda environment with Python >= 3.9: conda create -n subflow python=3.9
Activate the environment: conda activate subflow
Install subflow: pip install subflow
Follow the instructions to install Relion and cryoSPARC
Install crYOLO and pyem into their own anaconda environments
Download the multi-curve-fitting and filament subtraction scripts.
Edit the config.json file in the subflow installation to point to the correct crYOLO and pyem files, as well as the multi-curve-fitting and subtraction scripts

Start the GUI by running the following with the conda environment active:

subflow &

After saving parameters to a file (e.g. subflow.txt), re-opening the GUI at a later date opens the previous parameters. Saving is not automatic.

Tabs with hollow circles can be run on-the-fly (i.e. monitor files from the previous step). They turn into a black dot once they are running.

Clicking twice on a tab reveals a display tab to view the resulting micrographs for that step.

If Subflow crashes while Relion jobs are rerunning in the Preproc tab, they will continue to run. You can check this by running the Relion GUI. You may need to kill those jobs before being able to run Preproc again, or just continue in the Relion GUI.
Since cryosparc-tools is set to version 4.6.0 in the installation, your cryoSPARC installation must be 4.6.0. Otherwise cryosparc-tools needs to be downgraded or upgraded.

This project is licensed under the MIT License - see the LICENSE.txt file for details.