Walkthrough on membrane models - Revision history

Raffaele Coray at 12:43, 2 December 2024

2024-12-02T12:43:16Z

← Older revision		Revision as of 12:43, 2 December 2024
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			[[Category:Walkthroughs]]

	This walkthrough is a step-by-step guide that teaches how to extract subtomograms from structures with large surfaces (e.g., membranes) using the ''Dynamo'' surface models.		This walkthrough is a step-by-step guide that teaches how to extract subtomograms from structures with large surfaces (e.g., membranes) using the ''Dynamo'' surface models.

Stefano Scaramuzza: /* Generate subtomogram coordinates */

2021-08-13T08:23:30Z

Generate subtomogram coordinates

← Older revision		Revision as of 08:23, 13 August 2021
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	[[ File:membrane_step_17.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_17.png \|thumb\|center\| 700px\|]]

	~~Visualize~~ the crop positions:		Press the button ''create final positions and angles for particle cropping'' and visualize the crop positions:

	[[ File:membrane_step_18.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_18.png \|thumb\|center\| 700px\|]]

Stefano Scaramuzza at 15:13, 12 August 2021

2021-08-12T15:13:26Z

← Older revision		Revision as of 15:13, 12 August 2021
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	[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]

	These are 2D representations (projections) of the actually cropped 3D subtomograms. Thanks to our model, the particles are already oriented having the z axis normal to the surface and the x- and y-axis are inside the membrane surface. This is why the x-view of the particles shows a cut through the membrane:		These are 2D representations (projections) of the actually cropped 3D subtomograms. Thanks to our model, the particles are already oriented having the z axis normal to the surface, and the x- and y-axis are inside the membrane surface. This is why the x-view of the particles shows a cut through the membrane:

	[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]

Stefano Scaramuzza at 15:11, 12 August 2021

2021-08-12T15:11:27Z

Stefano Scaramuzza: /* Manual annotation */

2021-08-12T14:59:53Z

Manual annotation

← Older revision		Revision as of 14:59, 12 August 2021
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	We now ~~add~~ manually add model points along the surface of the membrane. To do that, first move the slice up to a high z height. Then place the mouse cursor on the membrane and press the key [c] on your keyboard. This adds a new model point. Continue adding model points as shown below. 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.		We now manually add model points along the surface of the membrane. To do that, first move the slice up to a high z height. Then place the mouse cursor on the membrane and press the key [c] on your keyboard. This adds a new model point. Continue adding model points as shown below. 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.

	[[ File:membrane_step_5.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_5.png \|thumb\|center\| 700px\|]]
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	[[ File:membrane_step_6.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_6.png \|thumb\|center\| 700px\|]]

	Repeat this procedure for the full structure until you get something similar to ~~the results shown below~~:		Repeat this procedure for the full structure until you get something similar to this:

	[[ File:membrane_step_7.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_7.png \|thumb\|center\| 700px\|]]

	We now need to define ~~where is~~ the "inside" of the structure. Move the slice somewhere in the middle, place your mouse cursor in the center of the structure and press the key combination of [shift]+[c]. A point with different color should appear~~, as shown below~~:		We now need to define the "inside" of the structure. This will be needed to create initial orientations of the subtomograms. Move the slice to somewhere in the middle, place your mouse cursor in the center of the structure and press the key combination [shift]+[c]. A point with a different color should appear:

	[[ File:membrane_step_8.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_8.png \|thumb\|center\| 700px\|]]

Stefano Scaramuzza at 14:55, 12 August 2021

2021-08-12T14:55:09Z

← Older revision		Revision as of 14:55, 12 August 2021
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	[[ File:membrane_step_13.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_13.png \|thumb\|center\| 700px\|]]

	To refine the depiction mesh change the parameter ''subdivision steps'' from 2 to 1. Press the button ''refine the depiction mesh'' once and visualize the result with the view button from the previous step. You should see a finer mesh. You should also see an increase in the number for ''# Depiction triangles'' below the previous button. Clicking the refinment button again, will further refine the mesh. If you want to go back to the original mesh, simply click the button of the previous step ''create a mesh for depiction'' to overwrite the current mesh.		To refine the depiction mesh change the parameter ''subdivision steps'' from 2 to 1. Press the button ''refine the depiction mesh'' once and visualize the result with the view button from the previous step. You should see a finer mesh. You should also see an increase in the number for ''# Depiction triangles'' below the previous button (in this case 1012). Clicking the refinment button again, will further refine the mesh. If you want to go back to the original mesh, simply click the button of the previous step ''create a mesh for depiction'' to overwrite the current mesh.

	[[ File:membrane_step_14.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_14.png \|thumb\|center\| 700px\|]]
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	[[ File:membrane_step_17.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_17.png \|thumb\|center\| 700px\|]]

	Visualize the crop positions as shown below. As you can see, each tile has one crop point in its center. In a real case you would try out different parameters until you have the coverage that fits your sample geometry. While setting those parameters, also always keep in mind the subtomogram box size that you plan to use.		Visualize the crop positions as shown below. As you can see, each tile has one crop point in its center. In a real case you would try out different parameters until you have the coverage that fits your sample geometry. While setting those parameters, also always keep in mind the subtomogram box size that you plan to use. The number next to ''# crop points'' (in this example 180) shows the total number of subvolumes that will be cropped:

	[[ File:membrane_step_18.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_18.png \|thumb\|center\| 700px\|]]

Stefano Scaramuzza at 14:52, 12 August 2021

2021-08-12T14:52:31Z

← Older revision		Revision as of 14:52, 12 August 2021
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	[[ File:membrane_step_17.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_17.png \|thumb\|center\| 700px\|]]

	Visualize the crop positions as shown below. As you can see, each tile has one crop point in its center. ~~Ina~~ real case you would try out different parameters until you have the coverage that fits your sample geometry. While setting those parameters, also always keep in mind the subtomogram box size that you plan to use.		Visualize the crop positions as shown below. As you can see, each tile has one crop point in its center. In a real case you would try out different parameters until you have the coverage that fits your sample geometry. While setting those parameters, also always keep in mind the subtomogram box size that you plan to use.

	[[ File:membrane_step_18.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_18.png \|thumb\|center\| 700px\|]]

	You can add the orientation of the particles to the plot as shown below. The shown orientation represents the z-axis of the particles taht will be cropped using this model.		You can add the orientation of the particles to the plot as shown below. The shown orientation represents the z-axis of the particles taht will be cropped using this model. The surface model as used here imposes the z-axis to be normal to the surface:

	[[ File:membrane_step_19.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_19.png \|thumb\|center\| 700px\|]]
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	[[ File:membrane_step_212.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_212.png \|thumb\|center\| 700px\|]]

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

	[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]

			These are 2D representations (projections) of the actually cropped 3D subtomograms. Thanks to our model, the particles are already oriented having the z axis normal to the surface and the x- and y-axis are inside the membrane surface. This is why the x-view of the particles shows a cut through the membrane:

	[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]

Stefano Scaramuzza at 14:45, 12 August 2021

2021-08-12T14:45:36Z

← Older revision		Revision as of 14:45, 12 August 2021
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	[[ File:membrane_step_21.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_21.png \|thumb\|center\| 700px\|]]

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

	[[ File:membrane_step_212.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_212.png \|thumb\|center\| 700px\|]]

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

	[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]


	[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]

Stefano Scaramuzza at 14:44, 12 August 2021

2021-08-12T14:44:23Z

← Older revision		Revision as of 14:44, 12 August 2021
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	[[ File:membrane_step_21.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_21.png \|thumb\|center\| 700px\|]]

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

			[[ File:membrane_step_212.png \|thumb\|center\| 700px\|]]


	[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]
	[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]

Stefano Scaramuzza at 14:43, 12 August 2021

2021-08-12T14:43:33Z

← Older revision		Revision as of 14:43, 12 August 2021
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	[[ File:membrane_step_16.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_16.png \|thumb\|center\| 700px\|]]

	The mesh can interpreted in a way that in the center of each tile (or "triangle") one coordinate for tomogram extraction will be placed. Refine the mesh in the same way as done before by ~~clocking~~ on ''refine the cropping mesh'':		The mesh can interpreted in a way that in the center of each tile (or "triangle") one coordinate for tomogram extraction will be placed. Refine the mesh in the same way as done before by clicking once on ''refine the cropping mesh'':

	[[ File:membrane_step_17.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_17.png \|thumb\|center\| 700px\|]]

			Visualize the crop positions as shown below. As you can see, each tile has one crop point in its center. Ina real case you would try out different parameters until you have the coverage that fits your sample geometry. While setting those parameters, also always keep in mind the subtomogram box size that you plan to use.

			[[ File:membrane_step_18.png \|thumb\|center\| 700px\|]]

			You can add the orientation of the particles to the plot as shown below. The shown orientation represents the z-axis of the particles taht will be cropped using this model.

	~~[[ File:membrane_step_18.png \|thumb\|center\| 700px\|]]~~
	[[ File:membrane_step_19.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_19.png \|thumb\|center\| 700px\|]]

			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:

	[[ File:membrane_step_20.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_20.png \|thumb\|center\| 700px\|]]

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

	[[ File:membrane_step_21.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_21.png \|thumb\|center\| 700px\|]]


	[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]
	[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]

← Older revision		Revision as of 14:45, 12 August 2021
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	[[ File:membrane_step_21.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_21.png \|thumb\|center\| 700px\|]]

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

	[[ File:membrane_step_212.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_212.png \|thumb\|center\| 700px\|]]

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

	[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]


	[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]

← Older revision		Revision as of 14:44, 12 August 2021
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	[[ File:membrane_step_21.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_21.png \|thumb\|center\| 700px\|]]

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

			[[ File:membrane_step_212.png \|thumb\|center\| 700px\|]]


	[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]
	[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]

← Older revision		Revision as of 14:43, 12 August 2021
Line 68:		Line 68:
	[[ File:membrane_step_16.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_16.png \|thumb\|center\| 700px\|]]

	The mesh can interpreted in a way that in the center of each tile (or "triangle") one coordinate for tomogram extraction will be placed. Refine the mesh in the same way as done before by ~~clocking~~ on ''refine the cropping mesh'':		The mesh can interpreted in a way that in the center of each tile (or "triangle") one coordinate for tomogram extraction will be placed. Refine the mesh in the same way as done before by clicking once on ''refine the cropping mesh'':

	[[ File:membrane_step_17.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_17.png \|thumb\|center\| 700px\|]]

			Visualize the crop positions as shown below. As you can see, each tile has one crop point in its center. Ina real case you would try out different parameters until you have the coverage that fits your sample geometry. While setting those parameters, also always keep in mind the subtomogram box size that you plan to use.

			[[ File:membrane_step_18.png \|thumb\|center\| 700px\|]]

			You can add the orientation of the particles to the plot as shown below. The shown orientation represents the z-axis of the particles taht will be cropped using this model.

	~~[[ File:membrane_step_18.png \|thumb\|center\| 700px\|]]~~
	[[ File:membrane_step_19.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_19.png \|thumb\|center\| 700px\|]]

			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:

	[[ File:membrane_step_20.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_20.png \|thumb\|center\| 700px\|]]

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

	[[ File:membrane_step_21.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_21.png \|thumb\|center\| 700px\|]]


	[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_22.png \|thumb\|center\| 700px\|]]
	[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_23.png \|thumb\|center\| 700px\|]]

← Older revision		Revision as of 15:11, 12 August 2021
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	[[ File:membrane_step_12.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_12.png \|thumb\|center\| 700px\|]]

	We now create two types of triangulations (or ''~~mesh~~''). One is for depiction only and the other will be used to define the coordinates at which the subtomograms will be extracted. Both meshes are created by first generating a coarse mesh and then refining it. To create the coarse mesh for depiction first click on the ''create a mesh for depiction'' button and visualize the results:		We now create two types of triangulations (or ''meshes''). One is for depiction only and the other will be used to define the coordinates at which the subtomograms will be extracted. Both meshes are created by first generating a coarse mesh and then refining it. To create the coarse mesh for depiction first click on the ''create a mesh for depiction'' button and visualize the results:

	[[ File:membrane_step_13.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_13.png \|thumb\|center\| 700px\|]]

	To refine the depiction mesh change the parameter ''subdivision steps'' from 2 to 1. Press the button ''refine the depiction mesh'' once and visualize the result with the view button from the previous step. You should see a finer mesh. You should also see an increase in the number for ''# Depiction triangles'' below the previous button (in this case 1012). Clicking the refinment button again, will further refine the mesh. If you want to go back to the original mesh, simply click the button of the previous step ''create a mesh for depiction'' to overwrite the current mesh.		To refine the depiction mesh change the parameter ''subdivision steps'' from 2 to 1. Press the button ''refine the depiction mesh'' once and visualize the result with the view button from the previous step. You should see a finer mesh. You should also see an increase in the number for ''# Depiction triangles'' below the previous button (in this case 1012). Clicking the refinment button again will further refine the mesh. If you want to go back to the original mesh, simply click the button of the previous step ''create a mesh for depiction'' to overwrite the current mesh.

	[[ File:membrane_step_14.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_14.png \|thumb\|center\| 700px\|]]
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	[[ File:membrane_step_16.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_16.png \|thumb\|center\| 700px\|]]

	The mesh can interpreted in a way that in the center of each tile (or "triangle") one coordinate for tomogram extraction will be placed. Refine the mesh in the same way as ~~done~~ before by clicking once on ''refine the cropping mesh'':		The mesh can be interpreted in a way that in the center of each tile (or "triangle"), one coordinate for tomogram extraction will be placed. Refine the mesh the same way as before by clicking once on ''refine the cropping mesh'':

	[[ File:membrane_step_17.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_17.png \|thumb\|center\| 700px\|]]

	Visualize the crop positions as shown below. As you can see, each tile has one crop point in its center. In a real case you would try out different parameters until you have the coverage that fits your sample geometry. While setting those parameters, also always keep in mind the subtomogram box size that you plan to use. The number next to ''# crop points'' (in this example 180) shows the total number of subvolumes that will be cropped:		Visualize the crop positions:

	[[ File:membrane_step_18.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_18.png \|thumb\|center\| 700px\|]]

	You can add the orientation of the particles to the plot as shown below. The shown ~~orientation represents~~ the z-axis of the particles ~~taht~~ will be ~~cropped using this~~ model. ~~The surface~~ model as ~~used here imposes~~ the z-axis to be normal to the surface:		As you can see, each tile has one crop point in its center. In a real case you would try out different parameters until you have the coverage that fits your sample geometry. While setting those parameters, also always keep in mind the subtomogram box size that you plan to use. The number next to ''# crop points'' (in this example 180) shows the total number of subvolumes that will be cropped. You can add the orientation of the particles to the plot as shown below. The shown orientations represent the z-axis of the particles that will be imposed by the model. Using the model as shown in this tutorial, the z-axis is set to be normal to the membrane surface:

	[[ File:membrane_step_19.png \|thumb\|center\| 700px\|]]		[[ File:membrane_step_19.png \|thumb\|center\| 700px\|]]