Hi Nancy,
In 1.4, normalization should include ramping background correction. Are
these images which you send? What format are they in? I cannot seem to
open them.
best,
Sjors
PS: shouldn't you run relion_preprocess_mpi instead of relion_preprocess?
(this will not solve your issue)
> Hi all,
>
> I'm concerned about whether a particle set is being normalized properly
> during particle extraction in Relion 1.4. Attached is a star file of
> representative particles (icosahedral virus). At the bottom is my particle
> extraction command.
>
> Backgrounds seem to vary quite a bit, even when boxes are not over the
> edge of the micrograph. I should say a good number of the micrographs show
> a regular light/dark gradient across the image (due to acquisition
> issues), and that each contain at least a single row of "dead" pixels.
> Particles were extracted from raw micrographs following CTF estimation in
> Relion using .box coord files determined in EMAN2, so these images should
> not have any normalization done to them except that specified by Relion's
> --norm option.
>
> Concerned that intensity variation will be problematic downstream -- can
> anyone suggest Relion-compatible methods to take into account the
> background gradients (if that's the culprit), or am I doing something more
> obviously wrong? Thanks for any input!
>
> Command for particle extraction:
> -bash-4.1$ srun --nodes 5 --ntasks-per-node 24 relion_preprocess --o ptcls
> --mic_star micrographs_ctf.star --coord_suffix ".box" --extract
> --extract_size 384 --norm --bg_radius 190 --white_dust -1 --black_dust -1
> --coord_files "Micrographs/*.box"
>
> -- particles themselves are approximately 260A diam
> -- also attached an example of early 2D classes showing background
> variation
>
> Thanks!!
>
--
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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