Re: [scipion-users] 2D masking particles

[Gregory S. <sha...@gm...>]

Oh, I missed the last part of your message. Scipion viewer by default
applies particle orientations when displaying, so you might not see the
difference between particles before and after running xmipp apply alignment
protocol.


Best regards,
Grigory

--------------------------------------------------------------------------------
Grigory Sharov, Ph.D.

MRC Laboratory of Molecular Biology,
Francis Crick Avenue,
Cambridge Biomedical Campus,
Cambridge CB2 0QH, UK.
tel. +44 (0) 1223 267542 <+44%201223%20267542>
e-mail: gs...@mr...

On Thu, Mar 1, 2018 at 5:21 PM, Gregory Sharov <sha...@gm...>
wrote:

> Hello Teige,
>
> there are a couple of things you should consider: 1) 2d classification in
> relion will always start from random angles unless you selected option
> "Consider previous alignment" (which in old code would set priors=angles);
> 2) before applying the mask you should use "xmipp-apply alignment 2d"
> protocol to produce a new stack of aligned particles. Alignment assign
> protocol will only create a "link" between particle set and aligments.
>
> So the workflow in your case would probably be: relion 3D autorefine ->
> relion2D without search (consider prev. alignment=yes) -> (output classes
> are therefore oriented as per the 3d map) -> take the side-view class and
> create a mask -> create particle subset from relion2D of side-view class ->
> xmipp3 apply alignment to subset --> xmipp3 apply 2d mask to the previous
> output
>
> Best regards,
> Grigory
>
> ------------------------------------------------------------
> --------------------
> Grigory Sharov, Ph.D.
>
> MRC Laboratory of Molecular Biology,
> Francis Crick Avenue,
> Cambridge Biomedical Campus,
> Cambridge CB2 0QH, UK.
> tel. +44 (0) 1223 267542 <+44%201223%20267542>
> e-mail: gs...@mr...
>
> On Thu, Mar 1, 2018 at 4:56 PM, Matthews-Palmer, Teige <
> t.m...@im...> wrote:
>
>> Dear Scipion Users,
>>
>> I have a situation where a domain of my object does not align in 2D or 3D
>> (some failed attempts of density subtraction & masked refinement).
>> I have side-views where this domain is separate (not overlapping) from
>> the rest of the object.
>> What I would like to do is to apply a fairly tight 2D mask around this
>> domain to the particle images. The xmipp programs in scipion seem to
>> provide a convenient way, however I cannot get it to work right yet.
>>
>> My workflow is: relion 3D autorefine -> relionc2D without search ->
>> (output classes are therefore oriented as per the 3d map) -> take the
>> side-view class and create a mask -> import mask -> create particle subset
>> from relionc2D of side-view class -> xmipp3 apply 2d mask
>> The outcome is that the mask is rotated in the output particles, but the
>> transformations are wrong (not even close). If I perform relionc2D without
>> search on the unmasked side-view particle subset, they do not recreate the
>> class average so they must not have the alignments assigned.
>>
>> If I do instead:
>> create particle subset from relionc2D of side-view class -> scipion
>> alignment assign using relion 3D autorefine as input alignments -> xmipp
>> apply 2d mask
>> Then, checking the particle subset, it can recreate the class average
>> without searching, so it has the alignments, but the mask is not rotated at
>> all in the output particles.
>>
>> So I tried xmipp3 apply alignment 2d to see if it will actually perform a
>> transformation and interpolation on the particle images and create a new
>> particle stack, but the output particles are unchanged.
>>
>> Does anyone have experience of 2D masking and have a workflow that works?
>> Any help greatly appreciated.
>> All the best,
>> Teige
>>
>>
>>
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>
>