After Refinement (Het. + Hom. + NU + Local) on cryosparc I get a map which cryosparc says is at an overall resolution of ~3.7A (with local resolution of 60-70% of the protein core (membrane domain) at a ~ 3.0-3.3A)
I take the same set of particles and take it through refinement on Relion and the post-processed maps looks better - I don’t see any more high resolution feature than that in the cryosparc maps but some of the relatively lower resolution domains are clearer and several of the beta-strands are also clearer in the Relion map. Nevertheless Relion says that its map is ~4.1A (with the core at ~3.5-3.8A).
I re-imported the Relion refined particles into cryosparc and did a local refinement job where I constrained the search space (angle: 1 in steps of 0.1 and offset: 0). The map looks improved than the prior cryosparc jobs reflecting the improved aspects from Relion refinement but the map resolution is ~3.7A.
I am not sure whether the ~0.4A difference in resolution is “real” so I’m a little unsure about what is the actual resolution here and how to proceed. Any advice here?
(The FSC curves of the cryosparc/relion reflect the difference in GSFSC resolution but there is nothing that is conspicuously wrong/suspicious in either of them (at least not to my eyes))
For post processing on Relion I generate the masks but on cryosparc I leave the mask as dynamic during the local refinement jobs. The mask generation parameters are identical on both Relion and cryosparc. As much as I can say on Chimera, the relion mask and the mask from the final iteration on cryosparc local refinement are very similar.
when you say the masks look identical, are you comparing with the refinement mask used in cryosparc or the fsc_auto mask? Just that often this can be the source of discrepancies, because csparc auto-shrinks the mask used for resolution determination
I was visually comparing the _volume_mask_refine.mrc with the Relion mask.
I just now looked at the _volume_mask_fsc and the _volume_mask_fsc_auto from the cryosparc refinement jobs and the _mask_fsc_auto is actually very tight/shrunk.
Cryosparc displays 2 sets of FSC curves, one labeled only as FSC and the FSC-mask auto-tightening. In my case, the first set, says my map is at a resolution of 3.93 and the second set (after auto tightening) says its at 3.78. So the _mask_fsc_auto is used for the 2nd set? What mask is used for resolution of the first set?
yes - I believe the fsc mask is used for the first one, and fsc_auto corresponds to the second. “Loose” corresponds to an FSC calculated with the refinement mask, which is what you were comparing with relion