Hi,
I think Giulia's suggestion is a good one. Sometimes at low-res the
orientations aren't well defined and starting from a high-res reference
may help. As always: just maintain your own critical judgement regarding
overfitting and final resolution estimates.
HTH,
Sjors
> Dear Gökhan,
>
> I had a similar issue. I didn't see any clear X-shaped artifacts but the
> map clearly wasn't right. The way I noticed was by running 3D
> classification from the refined particles, without allowing for
> alignments, and I got classes that were clearly the same molecule
> rotated. You might want to try this to see if this really is your problem.
>
> The way I solved it was by increasing the low-pass filter for the 3d
> refinements to 20A. Even a 40A filtered map was too "pseudo-2-fold" for
> the aligments to work, whereas the 20A-filtered one worked beautifully.
> You might need to find the coarser possible filter that eliminates the
> error - this will depend on the size and shape of your object.
>
> As long as the final resolution goes significantly beyond your initial
> filter I think you'll be fine - although I'd be very interested in
> hearing people's comments regarding this.
>
> Giulia
>
> On 28/03/2016 23:55, Gökhan Tolun wrote:
>>
>> Hi,
>>
>> I am working on two projects with no symmetry, and in the maps
>> generated by the refinement process I see artifacts that look as if
>> some of the particles were misaligned 180 degrees in one axis,
>> creating a pattern in the 3D reconstruction slices that looks like two
>> densities crossing each other (with a general X or + appearance at the
>> points where the two densities meet). Also, if I move through the
>> slices of the reconstructed 3d volume (e.g., using 3DMOD), the density
>> continuity moves in two different axes that are 90 degrees to each
>> other (best-effort explanation), so I am quite sure that there is
>> something wrong.
>>
>> I do not see any relevant parameters I can play with in the GUI, and
>> when I looked at the relion_refine parameters from the command line,
>> nothing obvious caught my eye that is listed as may be related to such
>> an issue.
>>
>> So, is there a way to avoid misalignment of particles during a C1
>> refinement? May any of the “expert” or other command-line parameters
>> be fiddled-with to fix such a faulty behavior during a 3D refinement,
>> and if so, which ones and how?
>>
>> Thanks.
>>
>> Gökhan
>>
>
> --
> Giulia Zanetti
> ISMB, Birkbeck College
> Malet St. London
> WC1E 7HX
> 02076316898
>
>
--
Sjors Scheres
MRC Laboratory of Molecular Biology
Francis Crick Avenue, Cambridge Biomedical Campus
Cambridge CB2 0QH, U.K.
tel: +44 (0)1223 267061
http://www2.mrc-lmb.cam.ac.uk/groups/scheres
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