Subset selection from particles stack



I imported my particle images after crYOLO picking and made 2D classifications. Then after the first refinement I subtracted one part of the structure and continued with local refinement. The end result (the aligned subtracted particles) was exported to a star file to make local 3D classifications in Relion without alignment. Now, after selecting the interesting class in Relion, I would like to do a new local refinement with a subset of particles in CryoSparc, but I don’t know how to do that.

I tried to import the subset star file of the subtracted particles, but I lose the (original CryoSparc) alignment parameters. Unfortunately, I don’t know how to use only a subset in CryoSparc or to relate the subtracted particle set to the original one to redo the full procedure from the beginning but only with the subset.

Any help would be appreciated.

Kind regards,



Hi @gabor,

When you imported the subset .star file back into cryoSPARC, did it not have the alignment parameters written out from Relion? If this is the case, the import of those particles should bring into cryoSPARC the alignments from Relion, which if I understand correctly, came from cryoSPARC when you exported the .star file. So you should have the “original cryoSPARC” alignment parameters when you import the subset, which would allow for local refinement in cryoSPARC.

Could you check the .star file from Relion’s subset to check that it has the needed alignment parameters?


Hi @apunjani,

The star file contains the alignment parameters, I double checked. I probably have my problem elsewhere. When I launch a local refinement with the subset particles I got the following error:
Traceback (most recent call last):

File “cryosparc2_worker/cryosparc2_compute/”, line 49, in
File “cryosparc2_compute/jobs/”, line 685, in check_default_inputs
assert False, 'Non-optional inputs from the following input groups and their slots are not connected: ’ + missing_inputs + ‘. Please connect all required inputs.’
AssertionError: Non-optional inputs from the following input groups and their slots are not connected: particles.alignments3D. Please connect all required inputs.

I don’t know what I miss here. How can I connect all required inputs?
Thank you in advance for your help!

Best regards,


Hi @gabor,

Take a look at the Job Builder Tutorial we made about using the job builder and connecting low-level results:

I’m not sure if this will fix your problem but hopefully it fills in some gaps