Segclassmodel

Program to assign a series of 3D models to class averages

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Input: Reference volumes

Output: Class average stack

Parameters

Parameter

Example (default)

Description

Reference volumes

recvol???.hdf

Input 3D reconstructions: to be reprojected and used for projection matching with provided classes. They should be generated using segmentrefine3d or segclassreconstruct because they require ‘rise/rotation’ and ‘point_sym’ entries in their header.

Class average stack

avgerages.hdf

Class average stack: accepted file formats (hdf).

spring.db file

spring.db

Program requires a combined spring.db from segment (and optional: micctfdetermine). An updated spring.db will be created in the working directory.

Pixel size in Angstrom

1.163

Pixel size is an imaging parameter (accepted values min=0.001, max=100).

Estimated helix width and height in Angstrom

(200, 600)

Generous width and height measure of helix required for rectangular mask (accepted values min=20, max=1500).

Sample parameter file

You may run the program in the command line by providing the parameters via a text file:

segclassmodel --f parameterfile.txt

Where the format of the parameters is:

Reference volumes                        = recvol???.hdf
Class average stack                      = avgerages.hdf
spring.db file                           = spring.db
Pixel size in Angstrom                   = 1.163
Estimated helix width and height in Angstrom = (200, 600)

Additional parameters (intermediate level)

Parameter

Example (default)

Description

Number of projections azimuthal/out-of-plane angle

(90, 7)

Number of projections (e.g. 90 azimuthal projections per 360 degrees - projection every four degrees, out-of-plane tilt 7 projections from -12 to 12 degrees to be searched for 3D reconstruction. High- and maximum resolution reconstructions will be further refined by 5 * number of specified azimuthal and out-of-plane angles (accepted values min=1, max=1000).

Out-of-plane tilt angle range

(-12, 12)

Expected out-of-plane tilt angle in degrees (0=no out-of-plane tilt) of helices used for 3D reconstruction (accepted values min=-40, max=40).

Keep intermediate files

False

Keep reprojection stack of models.

Limit in-plane rotation

True

Restricts in-plane rotation alignment.

Delta in-plane rotation angle

10.0

Rotational alignment is being performed within +/- delta angle from 0 and 180 degrees (delta=0 means only 0/180 are checked, delta=180 means no rotational restraint) (accepted values min=0, max=180).

X and Y translation range in Angstrom

(50, 50)

Translational range of alignment search perpendicular and along helix axis (X: off-center helical axis, Y: (minimum y-range=helical rise/2) (accepted values min=0, max=1000).

Internal binning factor

2

In order to speed up alignments images can be optionally binned (accepted values min=1, max=20).

High-pass filter option

False

Option to high-pass filter images before alignment or band-pass filter in combination with low-pass filter.

Low-pass filter option

True

Option to low-pass filter images before alignment or band-pass filter in combination with high-pass filter.

High and low-pass filter cutoffs in 1/Angstrom

(0.001, 0.09)

Filter design for high- and low-pass filter with cutoffs in 1/Angstrom. Maximum spatial frequency is 1/(2*pixelsize) (accepted values min=0, max=1).

MPI option

True

OpenMPI installed (mpirun).

Number of CPUs

8

Number of processors to be used (accepted values min=1, max=1000).

Temporary directory

/tmp

Temporary directory should have fast read and write access.

Sample parameter file (intermediate level)

You may run the program in the command line by providing the parameters via a text file:

segclassmodel --f parameterfile.txt

Where the format of the parameters is:

Reference volumes                        = recvol???.hdf
Class average stack                      = avgerages.hdf
spring.db file                           = spring.db
Number of projections azimuthal/out-of-plane angle = (90, 7)
Out-of-plane tilt angle range            = (-12, 12)
Keep intermediate files                  = False
Pixel size in Angstrom                   = 1.163
Estimated helix width and height in Angstrom = (200, 600)
Limit in-plane rotation                  = True
Delta in-plane rotation angle            = 10.0
X and Y translation range in Angstrom    = (50, 50)
Internal binning factor                  = 2
High-pass filter option                  = False
Low-pass filter option                   = True
High and low-pass filter cutoffs in 1/Angstrom = (0.001, 0.09)
MPI option                               = True
Number of CPUs                           = 8
Temporary directory                      = /tmp

Additional parameters (expert level)

Parameter

Example (default)

Description

Classes select option

False

Choose whether to select any particular classes.

Include or exclude classes

include

Choose whether to ‘include’ or ‘exclude’ specified classes.

Classes list

1-9, 11, 13

List of comma-separated class ids, e.g. ‘1-10, 12, 14’ (1st class is 0).

B-Factor

0

B-Factor in 1/Angstrom^2 to be applied to images. Zero no modulation of frequencies. Negative B-factors enhance high-resolution frequencies. Positive B-factors dampen high-resolution frequencies (accepted values min=-50000, max=50000).

Custom filter option

False

Option to custom filter images before alignment.

Custom-built filter file

filter_function.dat

Custom-built filter function with two columns (normalized spatial frequency 0 - 0.5, Fourier filter coefficients between 0 - 1): accepted file formats (txt, .dat).

Sample parameter file (expert level)

You may run the program in the command line by providing the parameters via a text file:

segclassmodel --f parameterfile.txt

Where the format of the parameters is:

Reference volumes                        = recvol???.hdf
Class average stack                      = avgerages.hdf
spring.db file                           = spring.db
Classes select option                    = False
Include or exclude classes               = include
Classes list                             = 1-9, 11, 13
Number of projections azimuthal/out-of-plane angle = (90, 7)
Out-of-plane tilt angle range            = (-12, 12)
Keep intermediate files                  = False
Pixel size in Angstrom                   = 1.163
Estimated helix width and height in Angstrom = (200, 600)
Limit in-plane rotation                  = True
Delta in-plane rotation angle            = 10.0
X and Y translation range in Angstrom    = (50, 50)
Internal binning factor                  = 2
High-pass filter option                  = False
Low-pass filter option                   = True
High and low-pass filter cutoffs in 1/Angstrom = (0.001, 0.09)
B-Factor                                 = 0
Custom filter option                     = False
Custom-built filter file                 = filter_function.dat
MPI option                               = True
Number of CPUs                           = 8
Temporary directory                      = /tmp

Command line options

When invoking segclassmodel, you may specify any of these options:

usage: segclassmodel [-h] [--g] [--p] [--f FILENAME] [--c] [--l LOGFILENAME] [--d DIRECTORY_NAME] [--version] [--classes_select_option]
                     [--keep_intermediate_files] [--limit_in-plane_rotation] [--high-pass_filter_option] [--low-pass_filter_option]
                     [--custom_filter_option] [--mpi_option]
                     [input_output [input_output ...]]

Program to assign a series of 3D models to class averages

positional arguments:
  input_output          Input and output files

optional arguments:
  -h, --help            show this help message and exit
  --g, --GUI            GUI option: read input parameters from GUI
  --p, --promptuser     Prompt user option: read input parameters from prompt
  --f FILENAME, --parameterfile FILENAME
                        File option: read input parameters from FILENAME
  --c, --cmd            Command line parameter option: read only boolean input parameters from command line and all other parameters will be assigned
                        from other sources
  --l LOGFILENAME, --logfile LOGFILENAME
                        Output logfile name as specified
  --d DIRECTORY_NAME, --directory DIRECTORY_NAME
                        Output directory name as specified
  --version             show program's version number and exit
  --classes_select_option, --cla
                        Choose whether to select any particular classes. (default: False)
  --keep_intermediate_files, --kee
                        Keep reprojection stack of models. (default: False)
  --limit_in-plane_rotation, --lim
                        Restricts in-plane rotation alignment. (default: False)
  --high-pass_filter_option, --hig
                        Option to high-pass filter images before alignment or band-pass filter in combination with low-pass filter. (default: False)
  --low-pass_filter_option, --low
                        Option to low-pass filter images before alignment or band-pass filter in combination with high-pass filter. (default: False)
  --custom_filter_option, --cus
                        Option to custom filter images before alignment. (default: False)
  --mpi_option, --mpi   OpenMPI installed (mpirun). (default: False)

Program flow

  1. prepare_models: Prepare provided models for 3D reprojection

  2. launch_segmentalign2d_to_match_classes_against_projections: Aligns class averages to model reprojections