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Specimen and experimental description


Specimen description

Specimen aggregation state:    
Specimen concentration (mg/mL): e.g. 1.35 (in mg/mL)
Buffer: e.g. 20mM NaCl, 10mM Tris-HCL,1mM MgCl2,1mM
Buffer pH: e.g. 7.4
Staining procedure: e.g. Grids with adsorbed protein floated on 1% w/v uranyl acetate for 20 seconds.
Specimen support details: e.g. 200 mesh gold grid with thin carbon support, glow discharged in amylamine atmosphere

Experiment description

Method:    
Imposed symmetry:   e.g. C1
Number of particles used in 3D reconstruction:   e.g. 3456
Number of class averages: e.g. 10
Image processing details: e.g. The particles were selected using an automatic selection program.

Specimen aggregation state

This should correspond to the physical state of the specimen when it is been imaged in the microscope.

Example: PARTICLE

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Concentration of the specimen (in mg/mL)

Give the concentration of macromolecular complexes in the sample.

Example: 7.4

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Buffer details of the specimen

Details of the constituents of the sample buffer: a solution containing either a weak acid and its salt or a weak base and its salt, which is resistant to changes in pH.Include names and concentration in terms of molarity or mass per unit volume.

Example: 25mM Hepes-KOH, 0.1 mM EDTA, 12.5 mM MgCl2, 10% (w/v) glycerol,100mM NaCl, 0.01% (v/v) Nonident P-40

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pH of the specimen

The pH value of the sample buffer.

Example: 7.4

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Staining procedure of the specimen

The quickest and most convenient way to examine samples is by electron microscopy of negatively stained specimens. All negative stains (uranyl acetate and sodium phosphotungstate are commonly used) are heavy metal compounds that form a glassy cast around macromolecular assemblies on drying. This cast is resistant to electron beam damage and is of high contrast, which makes direct observation of coarse structural features possible. However air-drying is necessary and the structure of biological specimens is conserved only to about 20 angstroms resolution and high resolution internal detail is lost. A non-vitrification preparation technique that maintains the specimen in its native, hydrated state when examined in the high vacuum of the electron microscope utilises the properties of thin films of glucose, trehalose and tannin, for example. The sugar compounds glucose and trehalose are rich in hydroxyl groups and so are able to mimic the native aqueous environment, although air-drying occurs, so these methods work best with fairly robust 2D crystals, such as purple membrane and phosphoporin, which are not damaged by air-drying. The most common preparation technique that maintains the specimen in its native,hydrated state when examined in the high vacuum of the electron microscope is that obtained by vitrification of the sample. Please enter your details on the 'Vitrification' page if this is the method that you have used.

Example: Grids with adsorbed protein floated on 1% w/v uranyl acetate for 20 seconds Details of the constituents of the sample buffer: a solution containing either a weak acid and its salt or a weak base and its salt, which is resistant to changes in pH.

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Details of specimen support

Details of the specimen support could include the type of support material, the size of grid mesh, the type of grid and any pretreatment of the grid and its supporting material.

Example: Holey carbon on top of 400 mesh gold grid.
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Crystal growth procedure

Details of the crystal growing methodology could include the type of apparatus used, the atmosphere, the temperature, the growth time, the number of crystals grown, and the mean crystal size.

Example: 2D-crystals were grown on a lipid monolayer.
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Crystal growth procedure

Details of the crystal growing methodology could include the type of apparatus used, the atmosphere, the temperature, the growth time, the number of crystals grown, and the mean crystal size.

Example: 2D-crystals were grown on a lipid monolayer.
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Crystal growth procedure

Details of the crystal growing methodology could include the type of apparatus used, the atmosphere, the temperature, the growth time, the number of crystals grown, and the mean crystal size.

Example: 2D-crystals were grown on a lipid monolayer.
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Plane/space Group

These are the 17 possible plane groups described using Herman Maugin nomenclature.

Example: P 3
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Unit Cell Length A

Unit cell length A in angstroms.

Example: 62.450
Range: 1.0 to 1000000.0
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Unit Cell Length B

Unit cell length B in angstroms.

Example: 62.450
Range: 1.0 to 1000000.0
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Enter Unit Cell Length C

Unit cell length C in angstroms.

Example: 62.450
Range: 1.0 to 1000000.0

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Unit cell angle alpha

Unit cell angle alpha in degrees.

Example: 120.00
Range: 0.1 to 179.9
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Unit cell angle beta

Unit cell angle beta in degrees.

Example: 120.00
Range: 0.1 to 179.9
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Unit cell angle gamma

Unit cell angle gamma in degrees.

Example: 120.00
Range: 0.1 to 179.9
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Crystal symmetry group

Example: C 2 2 21
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Unit Cell Length A

Unit cell length A in angstroms.

Example: 62.450
Range: 1.0 to 1000000.0
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Unit Cell Length B

Unit cell length B in angstroms.

Example: 62.450
Range: 1.0 to 1000000.0
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Enter Unit Cell Length C

Unit cell length C in angstroms.

Example: 62.450
Range: 1.0 to 1000000.0

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Unit cell angle alpha

Unit cell angle alpha in degrees.

Example: 120.00
Range: 0.1 to 179.9
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Unit cell angle beta

Unit cell angle beta in degrees.

Example: 120.00
Range: 0.1 to 179.9
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Unit cell angle gamma

Unit cell angle gamma in degrees.

Example: 120.00
Range: 0.1 to 179.9
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Helix handedness.

The handedness (sense) of the helix (screw operator). If you move along the helix ('walk through it') and it rotates in a clockwise fashion it is right handed.

Example: RIGHT HANDED
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Rotation per unit

The rotation angle per subunit.

Example: 20
Range: 1.0 to 180
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Translation per subunit.

The translation (rise) per subunit.

Example: 20
Range: 1.0 to 150
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Rotational symmetry.

The rotational symmetry of the helix. There are 2 allowed values: Cn or Dn, where n is the cyclic symmetry along the helical axis.

Helix handedness.

The handedness (sense) of the helix (screw operator). If you move along the helix ('walk through it') and it rotates in a clockwise fashion it is right handed.

Example: RIGHT HANDED
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Rotation per unit

The rotation angle per subunit.

Example: 20
Range: 1.0 to 180
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Translation per subunit.

The translation (rise) per subunit.

Example: 20
Range: 1.0 to 150
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Rotational symmetry.

The rotational symmetry of the helix. There are 2 allowed values: Cn or Dn, where n is the cyclic symmetry along the helical axis.

EM methodology

This should correspond to the primary experimental technique used for determining the structure which is being deposited.

Example: SINGLE PARTICLE RECONSTRUCTION

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Imposed symmetry

Point group symmetry applied to the deposited map, in Schoenflies notation (C[n-fold], D[n-fold], O, T, I), as described in 'Common conventions for interchange and archiving of three-dimensional electron microscopy information in structural biology.', J. Struct. Biol. 2005 (2):196-207. C1 is for asymmetric reconstruction.

Example: I
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Number of particles used in 3D reconstruction

The number of particles selected from the 2D projection images and used in the final 3D reconstruction.

Example: 3456
Range: 1 to 100000000
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Number of class averages

One class average contains images that are of the same 2D projection. Using multivariate statistical analysis of a sufficiently large number of good 2D projection images, and knowing the orientational relationship between all the projection class averages the 3D structure can be reconstructed. For an entirely asymmetric particle at least three different projections are required to solve the orientation problem.

Range: 1 to 1000000
Example: 30
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Image processing details

Additional details about the image processing related to single particles.

Example: The particles were selected interactively at the computer terminal.

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Imposed symmetry

Symmetry applied to the deposited map. If point group symmetry use Schoenflies notation (C[n-fold], D[n-fold], O, T, I), as described in 'Common conventions for interchange and archiving of three-dimensional electron microscopy information in structural biology.', J. Struct. Biol. 2005 (2):196-207. C1 is for asymmetric reconstruction.

Example: I
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Number of subtomograms averaged in 3D reconstruction

The number of subtomograms selected from tomogram(s) and used in the final 3D reconstruction.

Example: 25
Range: 1 to 5000
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Number of class averages

One class average contains subtomograms that are equivalent (v. gr. same conformation).

Range: 1 to 1000
Example: 30
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Image processing details

Additional details about the image processing related to subtomogram average.

Example: Subtomograms were selected manually.

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Imposed symmetry

Point group symmetry applied to the deposited map, in Schoenflies notation (C[n-fold], D[n-fold], O, T, I), as described in 'Common conventions for interchange and archiving of three-dimensional electron microscopy information in structural biology.', J. Struct. Biol. 2005 (2):196-207. C1 is for asymmetric reconstruction.

Example: I
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The number of projections used in the reconstruction of the tomographic map

The number of sections used in the reconstruction of the tomographic structure.
To reconstruct an object of 120 nm diameter with a resolution of 4 nm, the tilt increment has to be 2 degrees.
Due to the limited tilt range in the electron microscope (typically plus/minus 70 degrees), 70 projections have to be recorded.


Example: 70
Range: 0 to 1000
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Tomographic tilt angle increment (in degrees)

The tilt angle increment (in degrees) used in the reconstruction of the tomographic map. To reconstruct an object of 120 nm diameter with a resolution of 4 nm, the tilt increment has to be 2 degrees.

Example: 2
Range: 0.1 to 180 degrees
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Image processing details

Additional details about image processing related to tomographic data.

Example: CTF correction was performed on each projection.
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Image processing details

Additional details about image processing related to 2D crystals.

Example: Images were unbent using 2dx.
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Enter Other Details

Additional details about image processing related to helical particles.

Example: Particles were selected interactively at the computer terminal.
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