Bin micrographs

Cryo-EM Data Processing
17

  1. Import 3D Volumes

  2. Not sure what you mean - are you asking how to export to read into relion? Or another program? It will depend on what you want to do with them (in terms of how best to export)

18

Hi Oli,

Thanks for the 1st reply

for point no. 2, yes, - how to read the particles in another program - like say BSOFT or EMAN2 etc. I want to do some pre-processing for starting the model refinement or generate a starting model using BSOFT or EMAN 2. Can I get the particle images (stacks) from each micrograph (after frame alignment and dose correction).

Also can I get particles ctf corrected before the reconstruction process ?

19

Particle stacks will be located in the extract sub directory of the extraction job.

No, I believe particles are CTF corrected on the fly, so there are no stacks of CTF corrected particles.

20

HI Oli,

Continuing the thread here-

While carrying out class averaging for particles I encountered " illegal memory access was encountered". I do not find a solution to overcome this. Additionally this happened when I am trying to add another set of data to the existing one (which already provided a 3.4 Å resolution map).

Any feedback and a fix to the problem will be helpful.

Best
Andy

21

I’m not sure, but perhaps one of the cryosparc @team can help!

22

@Andy Please can you add more details about the error:

  • cryoSPARC job type
  • where in the logs or interface the error was encountered, with additional lines of output for context
  • the current “physical” box size of the particle stacks and any job parameters that affect the job’s memory complexity (resolution, number of classes, particles, etc.)
  • hardware and software:
  • how the other dataset was added to the existing one, and how the datasets differ
23

A post was split to a new topic: No particles from filament tracer

24

A post was merged into an existing topic: No particles from filament tracer

25

Hi Oli and wtempel,

The above issue also taken care of at present (BUT I will revisit it later to provide you more details). I have another problem.

  1. I recently completely a helical reconstruction with resolution up to 3.6Å. Now I want to generate the whole 3.6Å map to a much larger length (it is a phage tail density map). I will need the asymmetric map (at 3.6Å) so that I can symmetrize it to generate a 1200Å long map (the map is ~256Å long). How to obtain the asymmetric map ?

Is there any alternate way to do it ?

  1. I want to sort out heterogeneity based on the Pitch of the helix. I can generate maps for classifications from the asymmetric map, requested above. Following that How to I carry out the classification using the maps in cryosparc ?

Thanks

Andy

How do I do that?

26

Hi @Andy,
Thank you for posting these interesting questions.

To facilitate use and management of this forum, please post questions 1. and 2. separately as two new topics in the Helical Processing category.

27

Hi Andy,

Can I know how to bin micrographs in the CS? What’s the job name?

28

Hi

I binned the images in BSOFT (or you can do it in any package like EMAN2) and then imported them in Cryosparc.

However if memory serves well, there should be an option in cryosparc, I can possibly reply you back tomorrow or day after tomorrow. Some one from cryosparc team can definitely reply you, sooner but you can try the ‘Fourier crop to box size’.

Best

29

Depending on your stage of the workflow, “binning” can be accomplished in various job types.
For example, see this discussion about Patch Motion Correction.
Additionally, there are the Extract From Micrographs and Downsample Particles job types that can achieve a binning effect.

30

Hi Wtempel,

I’m running the Topaz Extract job, but it errored “out of memory”. I thought if I can bin the micrograph in a separate job, then do the Topaz extract, this error may cleared.

31

Hi @CleoShen - the defaults for topaz extract are quite memory hungry - you might try reducing the number of threads and processes, if you haven’t already.

Cheers
Oli

32

Hi Oli,

Thank you for your reply.
I binned the micrographs by Patch Motion Cor Job, then the Topaz Extract works, but I would like to downsample more, such as bin8, bin16, but the CS Patch Motion Cor F-crop option only provides as many as bin4, if you know how can I downsample more in CS?

33

This shouldn’t be necessary to run Topaz, as Topaz downsamples on the fly. Have you tried adjusting the downsampling factor in Topaz? What settings are you using?

34

Do you require down-sampling beyond factor 4 before picking, or would a downstream Downsample Particles job work for you?

35

@CleoShen
you may want to use some other package to downsample of desired value. What stage of processing you are in ?

36

@olibclarke @wtempel
I am taking this thread to ask about the helical processing questions that I have posted 2 months ago ? I think a separate thread is made but have not received any response. Can you help.