Dear Juraj,
Probably the input references are not on the same greyscale as the
reference projections. This could be because the latter did not come from
a reconstruction from the data themselves, or because the reconstruction
was done with different CTF settings (e.g. ignoring CTF until first peak).
You can use --first_iter_cc to avoid such problems.
HTH,
Sjors
> Dear all,
>
> I tried running a 2D classification using 2D projections from a 3D density
> as input references.
>
> The projections were generated with relion_project from the final output
> density from a Refine3D job, using 200 as number of projections. Then I
> ran the following command:
>
>
> `which relion_refine` \
> --ref projections_200/proj.mrcs \
> --ini_high 10 \
> --o supervised2D_v2/run \
> --i Refine3D/job150/run_ct15_data.star \
> --dont_combine_weights_via_disc \
> --pool 30 \
> --pad 2 \
> --ctf \
> --iter 1 \
> --tau2_fudge 2 \
> --particle_diameter 280 \
> --flatten_solvent \
> --zero_mask \
> --strict_highres_exp 10 \
> --oversampling 1 \
> --psi_step 10 \
> --offset_range 5 \
> --offset_step 2 \
> --norm \
> --scale \
> --j 2 \
> --gpu ""
>
> The result was as in the image attached - 200 classes which all look the
> same, as a smeared out almost circularly-symmetric density. In the
> data.star, all the particles have the "number of significant samples"
> value in the vicinity of 1300. The source projections themselves look as
> they ought to - different 2D projections, which correspond very well to
> the unsupervised 2D classes from which the 3D model was generated.
>
> [cid:92924e57-8e34-45d2-baad-6b1e3c8558ca]
>
> any hints as to what might be causing this / how to perform this
> correctly?
>
> Many thanks,
>
> Juraj
>
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--
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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