Recenter and rebox after subtraction

Local Refinement
2

Hi @rainfieldcn,

Thanks for the query – the recommended way to accomplish this in CryoSPARC is by performing the steps separately. For example, one possible workflow looks like:

EDIT: added two tweaks from @Hui (thanks!)

  1. Volume alignment tools (calculate mask center of mass):

    • Supply a volume, particles, and a mask covering the part of the map of interest; re-center to the center of this mask
    • This job will report the center of mass of the mask and can be used in the next job.

  2. Volume alignment tools (re-center):

    • Supply the same volume and particles as in job 1. Do not input any mask. Use the calculated center of mask as the 3D coordinates of new center.

  3. Extract from micrographs (re-extract)

    • Re-extract particles from Step 1 or 2 with “re-centering based on aligned shifts” enabled. Pick a box size large enough such that the particles are not cut off or too close to the edge. You can adjust the box size if it is too small after inspecting the output volume from step 4.

  4. Homogeneous reconstruction only

    • Use particles from step 3. Ensure that the output volume is not too close to the box edge

  5. Create a mask around the area to subtract from the output volume from Step 4

  6. Particle subtraction (subtract)

    • Connect the inputs below and run the job:
      • mask from Step 5,
      • particles from Step 3, and
      • volume from Step 4

  7. Downsample particles (crop)

    • Set the real-space crop size to a reasonable number, based on the desired final refinement.

Best,
Michael

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