Hi,
> For running a 3d refinement with partial signal subtracted images, can
> anyone suggest what works better as mask- spherical mask or a soft
> mask?
Soft mask.
> Also what reference should one use to ensure proper alignment of the
> signal subtracted particles?
The reference should contain only the volume that remains after subtraction.
The initial low pass filter might have to be weaker (say 10-15 A, instead of
40 A) but still at much lower resolution than the resolution of the region.
> As my area of interest is quite small compared
> to the whole particle, using a volume reference of only the focussed area
> simply isn't working. Although, running a recontsruction with the signal
> subtracted particle using relion_reconstruct program is giving a pretty
> good volume.
Did you also try local search?
How small is it? Even when it is too small to be aligned, you
might still be able to improve the density by running Class3D
without alignment to select a subset of particles that have
more-or-less the same relative orientation to the larger subtracted
region, and then refining the unsubtracted particles.
Best regards,
Takanori Nakane
> Thanks in advance!
>
> Shirin
>
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