PreProcessing: Difference between revisions
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As of version 1.1, the RELION GUI implements a semi-automated data preprocessing procedure. This procedures consists of three steps, each of which may be performed independently. The advantage of this procedure is ease-of-use (suitable STAR files and images are created with minimal user intervention) as well as speed (all | As of version 1.1, the RELION GUI implements a semi-automated data preprocessing procedure. This procedures consists of three steps, each of which may be performed independently. The advantage of this procedure is ease-of-use (''suitable STAR files and images are created with minimal user intervention'') as well as speed (''all time-consuming steps are MPI-parallelized''). | ||
== | = Filling in the GUI = | ||
To run the semi-automated preprocessing, select the <code>Preprocessing</code> run-type from the drop-down menu at the top of the GUI. | |||
The relion_preprocess program will launch the CTFFIND3 executable as indicated on the GUI | == I/O tab == | ||
Give a rootname for your output files and the general parameters of your microscope and detector setup. | |||
== CTFFIND tab == | |||
The relion_preprocess program will launch the CTFFIND3 executable as indicated on the GUI. Use wildcards (filenames with * or ?) to select multiple micrographs (in MRC format!) to work on, and then just provide all usual CTFFIND3 parameters. | |||
In some cases there are parts of the micrograph that are unsuitable for CTF estimation (e.g. labels on film). If that is the case, one may use the <code>Estimate CTF on window size (pix)</code> option to select the size of a squared window, placed at the center of the micrograph, that will be used for CTF-estimation. | In some cases there are parts of the micrograph that are unsuitable for CTF estimation (e.g. labels on film). If that is the case, one may use the <code>Estimate CTF on window size (pix)</code> option to select the size of a squared window, placed at the center of the micrograph, that will be used for CTF-estimation. | ||
=== | === extract tab === | ||
Again use wildcards (filenames with * or ?) to select all coordinate files for particle extraction, and then just provide a particle box size. | |||
=== operate tab === | |||
The extracted particles may be re-scaled, re-windowed, normalized, and have their contrast inverted (in that order). Keep re-scaled and re-windowed image sizes even numbers. '''Always normalize your particles''', and use a reasonable radius for the circle around your particles outside of which the standard deviation and average values for the noise are calculated. If there are white or black artefacts on the micrographs (e.g. caused by dust or hot/dead pixels), these may be removed by using a positive value for the dust removal options. All black/white pixels with values above the given parameter times the standard deviation of the noise are replaced by random values from a Gaussian distribution. For cryo-EM data, values around 3.5-5 are often useful. Make sure you do not erase part of the true signal. | |||
=== | === Running tab === | ||
The relion_preprocess program is MPI-parallelized: each MPI node will estimate CTF and extract particles for a subset of the selected micrographs/coordinate files. | |||
Revision as of 15:15, 11 September 2012
As of version 1.1, the RELION GUI implements a semi-automated data preprocessing procedure. This procedures consists of three steps, each of which may be performed independently. The advantage of this procedure is ease-of-use (suitable STAR files and images are created with minimal user intervention) as well as speed (all time-consuming steps are MPI-parallelized).
Filling in the GUI
To run the semi-automated preprocessing, select the Preprocessing run-type from the drop-down menu at the top of the GUI.
I/O tab
Give a rootname for your output files and the general parameters of your microscope and detector setup.
CTFFIND tab
The relion_preprocess program will launch the CTFFIND3 executable as indicated on the GUI. Use wildcards (filenames with * or ?) to select multiple micrographs (in MRC format!) to work on, and then just provide all usual CTFFIND3 parameters.
In some cases there are parts of the micrograph that are unsuitable for CTF estimation (e.g. labels on film). If that is the case, one may use the Estimate CTF on window size (pix) option to select the size of a squared window, placed at the center of the micrograph, that will be used for CTF-estimation.
extract tab
Again use wildcards (filenames with * or ?) to select all coordinate files for particle extraction, and then just provide a particle box size.
operate tab
The extracted particles may be re-scaled, re-windowed, normalized, and have their contrast inverted (in that order). Keep re-scaled and re-windowed image sizes even numbers. Always normalize your particles, and use a reasonable radius for the circle around your particles outside of which the standard deviation and average values for the noise are calculated. If there are white or black artefacts on the micrographs (e.g. caused by dust or hot/dead pixels), these may be removed by using a positive value for the dust removal options. All black/white pixels with values above the given parameter times the standard deviation of the noise are replaced by random values from a Gaussian distribution. For cryo-EM data, values around 3.5-5 are often useful. Make sure you do not erase part of the true signal.
Running tab
The relion_preprocess program is MPI-parallelized: each MPI node will estimate CTF and extract particles for a subset of the selected micrographs/coordinate files.