Thanks a lot! I returned to step 10 and remade an initial model excluding two bad classes instead of just one. I think it helped since now I could get ~6A on the second iteration of the second (->6A) refinement. Although I've got "No valid resolution found" for iterations 3 and 4 (see attachment), the result looks fine to me. Should I be worried? Best regards, Eugene Baulin On Sun, Aug 15, 2021 at 6:13 PM Ludtke, Steven J. wrote: > Usually this is caused by a bad initial model. With B-gal's symmetry and > shape, there are a couple of plausible but incorrect initial models which > can get things stuck in a local minimum for a while. This is generally > diagnosed at the initial model generation stage (step 10). Often it is > still possible to refine your way back to the correct structure with the > low resolution data, but it can take a dozen iterations or more to do it, > so it's usually better to go back to step 10 again. If you post some > screenshots of your refined map we can help diagnose if this is the issue. > > > > -------------------------------------------------------------------------------------- > Steven Ludtke, Ph.D. Baylor > College of Medicine > Charles C. Bell Jr., Professor of Structural Biology > Dept. of Biochemistry and Molecular Biology ( > www.bcm.edu/biochem) > Academic Director, CryoEM Core ( > cryoem.bcm.edu) > Co-Director CIBR Center ( > www.bcm.edu/research/cibr) > > > > > On Aug 15, 2021, at 8:35 AM, Eugene Baulin wrote: > > Hello! > > I'm totally new to the field of cryo-EM and now going through the EMAN2 > tutorial on single particle analysis with b-gal molecule ( > https://blake.bcm.edu/emanwiki/EMAN2/Tutorials?action=AttachFile&do=view&target=EMAN2_SingleParticleTutorial.pdf > ). > > I have a problem with the Refinement 2 step (step 12, part 2). > After the first refinement (with the aim of 15A), I was able to get 16.7, > 15.8, and 14.4A resolutions, based on a various number of input images (22, > 30, or all of them respectively). > But the second refinement (with the aim of 6A) gives me only ~9-10A > resolutions and, furthermore, the first iteration is always better in > resolution than the following (see the attached report, It took the 14.4A > structure as input). > > My EMAN2 version: > EMAN 2.91 final ( GITHUB: 2021-03-08 11:36 - commit: 81caed2 ) > Your Python version is: 3.7.7 > > What can I be doing wrong? > > Thanks in advance! > > Best regards, > Eugene Baulin > > > -- > -- > > ---------------------------------------------------------------------------------------------- > You received this message because you are subscribed to the Google > Groups "EMAN2" group. > To post to this group, send email to eman2@googlegroups.com > To unsubscribe from this group, send email to > eman2+unsubscribe@googlegroups.com > For more options, visit this group at > http://groups.google.com/group/eman2 > > --- > You received this message because you are subscribed to the Google Groups > "EMAN2" group. > To unsubscribe from this group and stop receiving emails from it, send an > email to eman2+unsubscribe@googlegroups.com. > To view this discussion on the web visit > https://groups.google.com/d/msgid/eman2/CAFtdTFXQq%3D%2BbqMnV7on5dmKwNcB5dHRB%2BrJKQm9Ussv%3DVLRiHw%40mail.gmail.com > > . > > > > -- > -- > > ---------------------------------------------------------------------------------------------- > You received this message because you are subscribed to the Google > Groups "EMAN2" group. > To post to this group, send email to eman2@googlegroups.com > To unsubscribe from this group, send email to > eman2+unsubscribe@googlegroups.com > For more options, visit this group at > http://groups.google.com/group/eman2 > > --- > You received this message because you are subscribed to the Google Groups > "EMAN2" group. > To unsubscribe from this group and stop receiving emails from it, send an > email to eman2+unsubscribe@googlegroups.com. > To view this discussion on the web visit > https://groups.google.com/d/msgid/eman2/C4716FB5-E846-4DF1-B554-54EFF207E581%40bcm.edu > > . >