.. _micctfdetermine: Micctfdetermine --------------- Program to determine CTF parameters from a set of micrographs using CTFFIND and CTFTILT (Mindell and Grigorieff JSB, 2003) +--------------------------------------------------+--------------------------------------------------+ | |micctfdetermine_input.png| | |micctfdetermine_output.pdf| | +--------------------------------------------------+--------------------------------------------------+ | Input: Micrographs | Output: Diagnostic plot pattern | +--------------------------------------------------+--------------------------------------------------+ .. |micctfdetermine_input.png| image:: ../gallery_images/micctfdetermine_input.png :alt: tree :width: 4 cm :target: ../_images/micctfdetermine_input.png .. |micctfdetermine_output.pdf| image:: ../gallery_images/micctfdetermine_output.pdf :alt: tree :width: 4 cm :target: ../_images/micctfdetermine_output.pdf .. _making_a_micctfdetermine_table_for_beginner: Parameters ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ .. tabularcolumns:: |p{4cm}|p{2.5cm}|p{9cm}| ============================================================================== ============================================================================== ==================================================================================================== Parameter Example (default) Description ============================================================================== ============================================================================== ==================================================================================================== Micrographs cs_scan034.tif Input micrographs: accepted file formats (tif, .mrc, .mrcs, .spi, .hdf, .img, .hed). Diagnostic plot pattern micctfdetermine_diag.pdf If single input micrograph: name of diagnostic plot file. In case of multiple input micrographs suffix to be attached to corresponding input micrograph. Output: accepted file formats (pdf, .png, .bmp, .emf, .eps, .gif, .jpeg, .jpg, .ps, .raw, .rgba, .svg, .svgz, .tif, .tiff). Spherical aberration 2.0 Spherical aberration - property of your electron microscope (accepted values min=0, max=10). Electron voltage in kV 200 Electron beam voltage of your microscope in kV (accepted values min=0, max=1000). Pixel size in Angstrom 1.163 Pixel size is an imaging parameter (accepted values min=0.001, max=100). ============================================================================== ============================================================================== ==================================================================================================== .. _micctfdetermine_parameter_file_for_beginner: Sample parameter file ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ You may run the program in the command line by providing the parameters via a text file:: micctfdetermine --f parameterfile.txt Where the format of the parameters is:: Micrographs = cs_scan034.tif Diagnostic plot pattern = micctfdetermine_diag.pdf Spherical aberration = 2.0 Electron voltage in kV = 200 Pixel size in Angstrom = 1.163 .. _making_a_micctfdetermine_table_for_intermediate: Additional parameters (intermediate level) ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ .. tabularcolumns:: |p{4cm}|p{2.5cm}|p{9cm}| ============================================================================== ============================================================================== ==================================================================================================== Parameter Example (default) Description ============================================================================== ============================================================================== ==================================================================================================== Range of defocus in Angstrom (10000, 45000) Images were taken at this defocus range in Angstrom. Positive values for underfocus. FEI microscopes tend to have an offset to the theoretical values of 6000 Angstrom (accepted values min=0, max=100000). Resolution search range in Angstrom (30.0, 6.0) Maximum and minimum resolution rings to be included in CTF determination (accepted values min=1, max=100). Binning option True Micrograph is reduced in size by binning. Binning factor 2 Micrograph is reduced in size by binning factor (accepted values min=1, max=20). MPI option True OpenMPI installed (mpirun). Number of CPUs 2 Number of processors to be used. Maximum number corresponds directly to number of input scans, i.e. no gain in performance if single input micrograph chosen (accepted values min=1, max=300). Temporary directory /tmp Temporary directory should have fast read and write access. ============================================================================== ============================================================================== ==================================================================================================== .. _micctfdetermine_parameter_file_for_intermediate: Sample parameter file (intermediate level) ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ You may run the program in the command line by providing the parameters via a text file:: micctfdetermine --f parameterfile.txt Where the format of the parameters is:: Micrographs = cs_scan034.tif Diagnostic plot pattern = micctfdetermine_diag.pdf Spherical aberration = 2.0 Electron voltage in kV = 200 Pixel size in Angstrom = 1.163 Range of defocus in Angstrom = (10000, 45000) Resolution search range in Angstrom = (30.0, 6.0) Binning option = True Binning factor = 2 MPI option = True Number of CPUs = 2 Temporary directory = /tmp .. _making_a_micctfdetermine_table_for_expert: Additional parameters (expert level) ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ .. tabularcolumns:: |p{4cm}|p{2.5cm}|p{9cm}| ============================================================================== ============================================================================== ==================================================================================================== Parameter Example (default) Description ============================================================================== ============================================================================== ==================================================================================================== Spring database option False If checked will read previous spring.db (Sqlite-compatible database) otherwise will create new one. spring.db file spring.db Program requires a previously generated spring.db and writes an updated spring.db database in the working directory. Continue spring.db option False Continue spring.db without re-computing previously determined CTF of micrographs. Do not tick in case you want to update the Ctf results of the micrographs in the provided database. Amplitude contrast 0.1 Amplitude contrast (between 0 and 1). For cryo data ranges from 0.07 to 0.14 and negative stain from 0.1 - 0.4 have been reported (accepted values min=0, max=1). Defocus search step size 300 Choose defocus search step size. Smaller step size will result in slower search (accepted values min=0, max=5000). Astigmatism search restraint in Angstrom 2000 Astigmatism in Angstrom. This is useful in case Thon rings are not well visible (accepted values min=0, max=50000). Tile size power spectrum in Angstrom 600 Tile size to be used for analysis. It will be internally converted to nearest power-of-two pixel dimensions (accepted values min=1, max=10000). CTFTILT refine option False Use CTFTILT to refine defocus only in case you tilted the stage. Local defocus search range 3000 Using the given search range for CTFTILT to refine locally around CTFFIND maximum (accepted values min=0, max=20000). Expected tilt and tilt search range in degrees (0, 8) Minimum and maximum micrograph tilt to be searched in CTFTILT determination (accepted values min=0, max=80). ============================================================================== ============================================================================== ==================================================================================================== .. _micctfdetermine_parameter_file_for_expert: Sample parameter file (expert level) ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ You may run the program in the command line by providing the parameters via a text file:: micctfdetermine --f parameterfile.txt Where the format of the parameters is:: Micrographs = cs_scan034.tif Diagnostic plot pattern = micctfdetermine_diag.pdf Spring database option = False spring.db file = spring.db Continue spring.db option = False Spherical aberration = 2.0 Electron voltage in kV = 200 Amplitude contrast = 0.1 Pixel size in Angstrom = 1.163 Range of defocus in Angstrom = (10000, 45000) Defocus search step size = 300 Astigmatism search restraint in Angstrom = 2000 Tile size power spectrum in Angstrom = 600 Resolution search range in Angstrom = (30.0, 6.0) CTFTILT refine option = False Local defocus search range = 3000 Expected tilt and tilt search range in degrees = (0, 8) Binning option = True Binning factor = 2 MPI option = True Number of CPUs = 2 Temporary directory = /tmp .. _using_micctfdetermine_on_cmdline: Command line options ~~~~~~~~~~~~~~~~~~~~~~~ When invoking micctfdetermine, you may specify any of these options:: usage: micctfdetermine [-h] [--g] [--p] [--f FILENAME] [--c] [--l LOGFILENAME] [--d DIRECTORY_NAME] [--version] [--spring_database_option] [--continue_spring.db_option] [--ctftilt_refine_option] [--binning_option] [--mpi_option] [input_output [input_output ...]] Program to determine CTF parameters from a set of micrographs using CTFFIND and CTFTILT (Mindell and Grigorieff JSB, 2003) 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 --spring_database_option, --spr If checked will read previous spring.db (Sqlite-compatible database) otherwise will create new one. (default: False) --continue_spring.db_option, --con Continue spring.db without re-computing previously determined CTF of micrographs. Do not tick in case you want to update the Ctf results of the micrographs in the provided database. (default: False) --ctftilt_refine_option, --ctf Use CTFTILT to refine defocus only in case you tilted the stage. (default: False) --binning_option, --bin Micrograph is reduced in size by binning. (default: False) --mpi_option, --mpi OpenMPI installed (mpirun). (default: False) Program flow ~~~~~~~~~~~~~~~~~~~~ #. prepare_micrograph_for_ctffind: Prepare micrographs for CTFFIND. #. run_ctffind_for_first_defocus_estimation: Runs CTFFIND on micrographs to determine defocus #. run_ctftilt_to_refine_defocus_parameters: Runs CTFTILT on micrographs to refine local defocus #. assemble_diagnostic_plot: Assembles summarized diagnostic plot of micrograph. Functions ~~~~~~~~~~~~~~~~~~~~ .. toctree:: :maxdepth: 1 micctfdetermine_functions