Difference between revisions of "Walkthrough on filament models"

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[[ File:filament_step_17.png |thumb|center| 700px|]]
 
[[ File:filament_step_17.png |thumb|center| 700px|]]
  
You get a new window with the model workflow. It has three large buttons (and input parameters below) that are executed sequentially starting from top. Click on ''create a smooth backbone'':
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You get a new window with the model workflow. It has three large buttons (and corresponding input parameters below) that are executed sequentially starting from top. Click on ''create a smooth backbone'':
  
 
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[[ File:filament_step_18.png |thumb|center| 500px|]]
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[[ File:filament_step_25.png |thumb|center| 500px|]]
 
[[ File:filament_step_25.png |thumb|center| 500px|]]
  
For the z and x axis you should see results similar to the following. These are 2D representations of the actually cropped 3D subtomograms:
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For the z and x axis you should see results similar to the following. These are 2D representations (projections) of the actually cropped 3D subtomograms:
  
 
[[ File:filament_step_26.png |thumb|center| 700px|]]
 
[[ File:filament_step_26.png |thumb|center| 700px|]]

Latest revision as of 15:52, 12 August 2021

This walkthrough is a step-by-step guide that teaches how to extract subtomograms from filament-like structures using the Dynamo filament models.

Data

We are going to demonstrate the basic ideas and tools on a synthetic dataset. This data is already available in the catalogue manager. It can be accessed by first opening the catalogue manager using the command:

dcm

After the catalogue manager opens, we create the synthetic tomogram that includes tubular looking objects (filaments) in the following way:

Filament step 1.png

A new tomogram entry appears in the catalogue manager. Any further annotations (models) to the tomogram will be linked to this entry. Select the entry and open it:

Filament step 2.png

A tomogram viewer opens. You might need to adjust the contrast (blue arrow). To move through the tomogram slices, you can either use the mouse wheel, click and drag the tomogram slice up and down (orange arrows), or move the position control left and right (orange box).

Filament step 3.png

Manual annotation

To be able to later annotate a filament of interest, we first create an alternative view by using the anchor points. We start by marking the beginning of a filament. Move the slice up to a high z height. Place the mouse in the center of a filament and press the key [1] on the keyboard to place an anchor in the beginning of the filament. Note: to correct the anchor point just click the [1] key again.

Filament step 4.png

To mark the end of the filament, move to a low z-height. Place the mouse in the center of the same filament and press the key [2] on the keyboard to place the anchor in the end of the filament.

Filament step 5.png

Finally, a third anchor point (key [3]) is placed to define the width of the alternative view that we are creating:

Filament step 6.png

Click on set plane (orange box) to define a plane along the anchor points:

Filament step 7.png

Use the anchor for plane selection control (orange box) to activate and rotate the viewing plane around the filament:

Filament step 8.png

Annotations are done within a so called model. We open a new model of type filament as follows:

Filament step 9.png

If necessary, click and drag while holding the control key to rotate the view of the tomogram to get a good view of the filament:

Filament step 10.png

To add annotation points to the model, place the mouse on the filament and press the key [c]. Use the right-click to delete points if needed. Note: A known bug may cause the clicked points to be generated slightly displaced from the mouse cursor. Maximize the window of the tomogram viewer to get rid of this effect. Generate about 10 points:

Filament step 11.png

We show you an alternative way to click the points on the filament. To demonstrate this, we first delete the current points as shown below:

Filament step 12.png

We create a new model in the same way as before:

Filament step 9.png

For this alternative method, we first create a view of orthogonal slices through the filament. In tools for filaments set sidelength to 64 and thickness to 10 (see orange box). Then click on extract (see orange box):

Filament step 13.png

You get a new window with the view of the orthogonal slices through the filament:

Filament step 14.png

Same as before, using the [c] key, add model points at the center of the tube filament:

Filament step 15.png

When done, close the window and you should see your clicked points in the main view:

Filament step 16.png

Generate subtomogram coordinates

Using the clicked model we can now define the coordinates where the subvolumes will be extracted. To do that, we first processes the clicked model points using the model workflow to generate a user specific cropping geometry. Open the workflow:

Filament step 17.png

You get a new window with the model workflow. It has three large buttons (and corresponding input parameters below) that are executed sequentially starting from top. Click on create a smooth backbone:

Filament step 18.png

This creates an interpolation through the points in the model. Visualize it by left-clicking on view:

Filament step 19.png

Add the clicked model points to the graph:

Filament step 20.png

This looks good. We now create coordinates along this interpolation (backbone) which will be used as centers for the subtomogram extraction (cropping). Click on create final positions and angles for particle cropping. Ignore the angles, as they are not part of this model. Visualize the crop positions:

Filament step 21.png

You can see that these are too many crop points. Adjust the spacing between the cropping points by changing the parameter subunits dz from 2 to 20 (see orange box). Click again create final positions and angles for particle cropping to update the information and visualize the new crop points:

Filament step 22.png

This looks much better. Save the model by clicking on save resulting model into catalogue and we can move to the subtomogram extraction.

Extract subtomograms

In this final step we extract subvolumes at the coordinates defined before. First, open the partcile extraction GUI:

Filament step 23.png

Set the sidelength to 64 and click on start cropping (orange box):

Filament step 24.png

To see the results, click on ddbrowse (orange box):


Filament step 245.png


In the new window activate the table, select the axis and click on show (orange boxes):

Filament step 25.png

For the z and x axis you should see results similar to the following. These are 2D representations (projections) of the actually cropped 3D subtomograms:

Filament step 26.png