[3dem] Relion Polishing with EER-movies containing varying frames and fractions
Takanori Nakane
tnakane.protein at osaka-u.ac.jp
Mon Apr 3 06:22:03 PDT 2023
Hi,
> Therefore, each subset has a different amount of EER fractions ( in the range of 600 – 1200 fractions).
Did you run 10 motion correction jobs with different EER groupings and
then join micrograph STAR files?
> Is this a bug in relion?
Probably yes. The program should read EER grouping for every movie but
somehow this is not working.
> How can I overcome this?
>
> Is the only way to first make movies with 50 frames from each subset and not work on the original EERs anymore? I processed fine to about 4 Angstroem
> without doing so, is it really necessary just for polishing to now make 50 frame movies, or work on each subset separately? (As far as I understand
> relion, it does not have a GUI-function to generate 50 frame movies from EER data, but it has a command-line function to do so, am I right?
Run Polish with the same run_data.star and postprocess.star but with
each micrograph STAR files BEFORE joining.
Polish with process only particles on the given micrographs.
Thus, after a chain of 10 Polish jobs, all particles will be polished.
Best regards,
Takanori Nakane
On 2023/04/03 21:56, Klink, Björn wrote:
> Hi there
>
> I encountered a weird problem with Relion 4 polishing in a dataset with 10 subsets of data from a Falcon 4i camera (EER movies).
>
> Each subset was measured with the same total exposure of 50 e-/A^2, but the exposure time per micrograph to achieve that was significantly different
> in some subsets.
>
> Therefore, each subset has a different amount of EER fractions ( in the range of 600 – 1200 fractions).
>
> Since in relion one can only choose how many EER fractions are combined to one frame, the amount of frames after EER fraction combination varies from
> 49 to 51.
>
> I figured out that with the –find-shortest option, a polishing job on all the datasets combined would find out that 49 is the amount of frames that is
> present in all micrographs, and I assume on datasets which are not in the EER format, everything would work out fine from here on.
>
> However, since the amount of fractions of the original EERs is still different, I found that the program crashes with the error message like:
>
> EERRenderer::renderFrames(frame_start = 967, frame_end = 987), NFrames = 918
>
> EERRenderer::renderFrames(frame_start = 904, frame_end = 924), NFrames = 918
>
> n:
> /Applic.HPC/Easybuild/zen3/2021a/build/RELION/4.0/foss-2021a-beta-543db0a-CUDA-11.3.1/relion-543db0a84a6735cc975be93fbafc0ffe683ebc57/src/renderEER.cpp, line 315
>
> ERROR:
>
> Invalid frame range was requested.
>
> I found that when I limit the amount of fractions to sth like 20, relion would not crash (maybe because then the highest amount of fractions in one of
> the datasets that contributed to the first 20 frames is not lower than the total amount of fractions in another subset). However, if 30 frames from
> one dataset are made from e.g. 750 fractions, while in the other subset the total 50 frames only were made from 690 fractions (because of higher
> exposure per second), the program would crash with above error.
>
> Is this a bug in relion?
>
> How can I overcome this?
>
> Is the only way to first make movies with 50 frames from each subset and not work on the original EERs anymore? I processed fine to about 4 Angstroem
> without doing so, is it really necessary just for polishing to now make 50 frame movies, or work on each subset separately? (As far as I understand
> relion, it does not have a GUI-function to generate 50 frame movies from EER data, but it has a command-line function to do so, am I right?
>
> Best wishes
>
> Björn
>
>
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