Dear users,
Sorry to piggy back on this thread but it seems like the most relevant thing I have come across to my question! I also have a 20nm D12 object where I am trying to do focussed classification and symmetry expansion to look for differences between monomers. I symmetry expand following auto-refine in Relion and signal subtract everything except the masked region of interest, using a quick relion_reconstruct to check that this has worked. Relion 3.1 offers the option to subtract centering on the mask of the area you want to keep and with a new (smaller) box size, but the problem is I would need to do the same with the mrc reference volume to run the classification. My question is, would this workflow make sense? I would not be re-aligning the images, so to me it does not make sense to work with large box sizes that are mostly subtracted signal, but perhaps I am missing something fundamental here? If it does make sense, could you point me to the pyem code? Thankyou very much for your help, Rhian