APPL_PRM is the second step in EM reconstruction after the alignment stage. With alignment parameters and the conversion factors for electron counts to mass density (both are loaded from the file pointed to by the input parameter IprmFile), this program creates a mass normalized and aligned data stack for reconstruction.
The algorithm is:
Here's an example command file for APPL_PRM:
(time appl_prm \
/mama/weiping/test/centSr.stk \
/mama/weiping/test/centSr.MnAln \
-iprmfile=/mama/weiping/test/centSr.bprmMn \
-dimxy=480:480 -iv=0:22:1 -shxyz=0:0:0 -iref=-1 \
-imform=0 -pcbase=.5 -positivity=1 ) \
> /mama/weiping/emrecon.log
IdatFile | OdatFile | NX:NY:NV | IprmFile | dimxy | iv | shXYZ | iRef | imForm | pcBase | positivity | tilt_offset
Priism | Reconstruction | MASSNORM | EWBP | TAPIR | Alignment
This is the name of the file containing the raw projection data stack from the CCD (i.e. measured in terms of electron counts; data stacks with the contrast inverted can not be handled).
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This parameter names the file that will contain the aligned and mass normalized data stack.
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The first two values are, respectively, the x and y dimension of the projections. The third value is the number of projections in the data stack.
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This parameter supplies the name of the file with the alignment information and, optionally, conversion factors for mass normalization. The file can be generated by BALIGN or by MASSNORM.
On the command line, use -iprmfile=filename
to supply the name of the parameter file. If the option is not set or
the filename is not supplied or is none, the program will
terminate with an error.
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These two values are, respectively, the x and y dimensions of the aligned projections. Usually the full dimensions of the input data are used, but when doing a quick reconstruction to determine an appropriate value for the z shift, it is convenient to generate a aligned projection with a y dimension of one.
On the command line, set the dimensions of the output data stack with
-dimxy=nx:ny. If not set,
the output x and y dimensions are the same as the x and y dimensions of the
input data stack.
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These three values set which of the input projections are aligned and mass normalized. The first value is the index of the first projection to use, the second value is the index of the last projection to use, and the third is the index step. Allowable values for the indices are between zero and the number of input projections minus one, inclusive.
From the command line, use
-iv=first:last:step to set the range of projections to process.
When not set, the default is to process the entire input stack: first is
zero, last is the number of input projections minus one, and
step is one.
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These three values (thery are in units of pixels) shift the imaginary 3D tilt axis of the projection data from its default location; they are used to center the reconstruction volume on an object of interest. The default orientation of the 3D tilt axis has it pass through the point whose x and y coordinates are the center of the reference projection (see the iRef parameter) and whose z coordinate is the z coordinate of the center of mass of the reference markers used for alignment.
A good way to determine appropriate values for the shifts is to use the Pick XYZ toggle buttons in the main reconstruction dialog.
Use -shxyz=x_shift:y_shift:z_shift
to set the shifts on the command line. If the shifts are not specified, a
value of zero pixels for each shift is assumed.
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For calculating the effect of the x and y shifts (see the shXYZ input parameter), the center of the projection whose index is given by iRef is used.
The reference projection can be set on the command line with
-iref=index. If it is not specified or
index is less than zero or greater than or equal to the
number of projections in the input file, the projection whose tilt
angle is closest to zero is used.
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This parameter sets which model should be assumed for the process of image formation - how electron counts are related to the mass density of the specimen. The three options are:
For more details on the image formation model, see the overview.
On the command line, use -imform=0 to use the log
conversion, -imform=1 to use the scaled linear model,
or -imform=2 to use the linear model. If none of them
are set, the log conversion is used.
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The mass normalization process (see the overview) includes an additive term which would correspond to the presence of a sheet with uniform mass density. The pcBase parameter scales the contribution of this term which has an important effect on positivity constraints. If pcBase is too small, a larger fraction of the mass normalized data will be negative and will be coerced to zero when a positivity constraint is applied; this coercion causes errors. If pcBase is too large, the mass normalized values will be positive and much greater than zero so a positivity constraint will provide little leverage for refining the reconstruction. As a compromise, the recommended value for pcBase is 0.5 if the Fit: background option to MASSNORM was used. When the Fit: average option is used with MASSNORM, a value of 1.0 is recommended for pcBase.
On the command line, set the scale factor with
-pcbase=factor. When it is not set, a value of 1.0
is assumed.
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When on, mass normalized values which are less than zero are coerced
to be zero. On the command line, this behavior is selected by specifying
the -positivity=any value option. By default,
if no such option is supplied on the command line, negative mass normalized
values are not coerced to be zero.
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The value of this parameter, assumed to be in degrees, is added to
the tilt angles stored in the header of the output data stack. It does
not affect the tilt angles that are used in the mass normalization
calculation. On the command line you can supply the tilt offset by specifying
-tilt_offset=angle. When no offset is set, a value
of zero is assumed.
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