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Dear Pranav, generally speaking, not about Relion because I don't know exactly how the mask is used, I don't see any problem in reconstructing without mask, then calculating a mask (making sure you don't leave out the object to reconstruct, and then redoing the alignment. The mask is used in this case to avoid the alignment to try to fit volume artifacts with some funny shape from the images. I like to call this "removing noise anchors from the volume". This mask does not necessarily be used when reconstructing (which might artificially increase the resolution because it is correlating Fourier coefficients). This is the strategy followed in Xmipp highres (accessible from Scipion). You may use a mask to align (to remove noise anchors) or no mask at all, and the same for the reconstruction (and the mask for the reconstruction may not be the same as the one for the alignment). Kind regards, Carlos Oscar El 08/02/2019 a las 18:03, Pranav Shah escribió: > Dear Cryo-EMers, > > I recently had a an idea regarding masking and 3d refinement in RELION > and was interested to know what the community thought about it. Some > background, my dataset consists of icosahedral viral particles whose > initial 3d refinement converged to 4.5A using default values and a > mask with an outer radius of 410A. Post -refinement I sharpened the > mask using a mask by setting the binarisation threshold value to an > appropriate level and with a soft edge of 5px. I then proceeded to run > CTFRefine with beam tilt estimation turned on. Now the question I have > is, is it kosher for me to run a 3d refinement of the ctfrefined > particle stack with the mask that was obtained in the initial Post > Process step? The way I see it is that, the mask was generated from a > map that was obtained in an unbiased manner, encloses just the capsid > and ignores the genome based density. I now want the program to refine > the parameters only in that region, and therefore feel justified in > using the mask used for sharpening to run a second round of > refinement. Is this fair? > > Looking forward to hearing your thoughts. > > Cheers, > Pranav > -- > Pranav Shah > Postdoctoral Research Fellow. > > Hogle Lab > Department of Biological Chemistry and Molecular Pharmacology, > Building C2 - First Floor > 240 Longwood Avenue > Boston, MA 02115 > (617) 432-4360 (fax) > (617) 432-3839 (lab) > > ------------------------------------------------------------------------ > > To unsubscribe from the CCPEM list, click the following link: > https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCPEM&A=1 > -- ------------------------------------------------------------------------ Carlos Oscar Sánchez Sorzano e-mail: [log in to unmask] Biocomputing unit http://i2pc.es/coss National Center of Biotechnology (CSIC) c/Darwin, 3 Campus Universidad Autónoma (Cantoblanco) Tlf: 34-91-585 4510 28049 MADRID (SPAIN) Fax: 34-91-585 4506 ------------------------------------------------------------------------ ######################################################################## To unsubscribe from the CCPEM list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCPEM&A=1