subtom_b_factor_by_subsets

Estimates the B-Factor by determine the resolution of subsets of particles as described in Rosenthal, Henderson 2003.

This subtomogram averaging analysis script uses three MATLAB compiled scripts below:

• subtom_maskcorrected_FSC_bfactor

• subtom_parallel_sums_bfactor

• subtom_weighted_average_bfactor

Options

Directories

scratch_dir

Absolute path to the folder with the input to be processed. Other paths are relative to this one.

mcr_cache_dir

Absolute path to MCR directory for the processing.

exec_dir

Directory for executables

Variables

sum_exec

Parallel Summing executable

avg_exec

Weighted Averaging executable

fsc_exec

FSC executable

Memory Options

mem_free

The amount of memory the job requires for alignment. This variable determines whether a number of CPUs will be requested to be dedicated for each job. At 24G, one half of the CPUs on a node will be dedicated for each of the processes (12 CPUs). At 48G, all of the CPUs on the node will be dedicated for each of the processes (24 CPUs).

mem_max

The upper bound on the amount of memory the alignment job is allowed to use. If any of the processes request or require more memory than this, the queue will kill the process. This is more of an option for safety of the cluster to prevent the user from crashing the cluster requesting too much memory.

Other Cluster Options

job_name

The job name prefix that will be used for the cluster submission scripts, log files, and error logs for the processing. Be careful that this name is unique because previous submission scripts, logs, and error logs with the same job name prefix will be overwritten in the case of a name collision.

array_max

The maximum number of jobs per cluster submission script. Cluster submission scripts work using the array feature common to queuing systems, and this value is the maximum array size used in a script. If the user requests more batches of processing than this value, then the submission scripts will be split into files of up to array_max jobs.

max_jobs

The maximum number of jobs for alignment. If the number of batches / exceeds this value the script will immediately quit.

run_local

If the user wants to skip the cluster and run the job locally, this value should be set to 1.

Subtomogram Averaging Workflow Options

Parallelization Options

iteration

The index of the reference to generate : input will be all_motl_fn_prefix_iteration.em (define as integer)

num_avg_batch

The number of batches to split the parallel subtomogram averaging job into.

File Options

all_motl_a_fn_prefix

Relative path and name prefix of the concatenated motivelist of all particles in the first half-map.

all_motl_b_fn_prefix

Relative path and name prefix of the concatenated motivelist of all particles in the second half-map.

ref_a_fn_prefix

Relative path and name prefix of the reference volumes of the first half-map.

ref_b_fn_prefix

Relative path and name prefix of the reference volumes of the second half-map.

ptcl_a_fn_prefix

Relative path and name prefix of the subtomograms that comprise the first half-map.

ptcl_b_fn_prefix

Relative path and name prefix of the subtomograms that comprise the second half-map.

weight_a_fn_prefix

Relative path and name prefix of the weight files for the first half-map.

weight_b_fn_prefix

Relative path and name prefix of the weight files for the second half-map.

weight_sum_a_fn_prefix

Relative path and name prefix of the partial weight files of the first half-map.

weight_sum_b_fn_prefix

Relative path and name prefix of the partial weight files of the second half-map.

output_fn_prefix

Relative path and prefix for the name of the output maps and figures.

Averaging Options

tomo_row

Which row in the motl file contains the correct tomogram number. Usually row 5 and 7 both correspond to the correct value and can be used interchangeably, but there are instances when 5 contains a sequential ordered value starting from 1, while 7 contains the correct corresponding tomogram.

iclass

Particles with that number in position 20 of motivelist will be added to new average (define as integer e.g. iclass=1). NOTES: Class 1 is ALWAYS added. Negative classes and class 2 are never added.

Mask Corrected FSC Workflow Options

File Options

fsc_mask_fn

Relative or absolute path and name of the FSC mask.

filter_a_fn

Relative or absolute path and name of the Fourier filter volume for the first half-map. If not using the option do_reweight just leave this set to “”

filter_b_fn

Relative or absolute path and name of the Fourier filter volume for the second half-map. If not using the option do_reweight just leave this set to “”

FSC Options

pixelsize

Pixelsize of the half-maps in Angstroms

nfold

Symmetry to applied the half-maps before calculating FSC (1 is no symmetry)

rand_threshold

The Fourier pixel at which phase-randomization begins is set automatically to the point where the unmasked FSC falls below this threshold.

plot_fsc

Plot the FSC curves - 1 = yes, 0 = no

Sharpening Options

do_sharpen

Set to 1 to sharpen map or 0 to skip and just calculate the FSC

box_gaussian

To remove some of the edge-artifacts associated with map-sharpening the edges of the map can be smoothed with a gaussian. Set to 0 to not smooth the edges, otherwise it must be set to an odd number. If an even number is given one will be added to the value to make it odd.

filter_mode

There are two mode used for low pass filtering. The first uses an FSC based threshold (mode 1), i.e. after FSC < 0.143, or a pixel-based resolution threhsold (mode 2).

filter_threshold

Set the threshold for the low pass filtering described above. Should be less than 1 for FSC based threshold (mode 1), and an integer value for the Fourier pixel-based threshold (mode 2).

plot_sharpen

Plot the sharpening curve - 1 = yes, 0 = no

Reweighting Options

do_reweight

Set to 1 to apply the externally calculated Fourier weights filter_A_fn and filter_B_fn to each half-map to reweight the final output map.

Example

scratch_dir="${PWD}"

mcr_cache_dir="${scratch_dir}/mcr"

exec_dir="/net/dstore2/teraraid/dmorado/software/subTOM/bin"

sum_exec="${exec_dir}/alignment/subtom_parallel_sums_bfactor"

avg_exec="${exec_dir}/alignment/subtom_weighted_average_bfactor"

fsc_exec="${exec_dir}/analysis/b_factor_by_subsets/subtom_maskcorrected_fsc_bfactor"

mem_free="1G"

mem_max="64G"

job_name="subTOM"

array_max="1000"

max_jobs="4000"

run_local="0"

iteration="1"

num_avg_batch="1"

all_motl_a_fn_prefix="even/combinedmotl/allmotl"

all_motl_b_fn_prefix="odd/combinedmotl/allmotl"

ref_a_fn_prefix="FSC/ref_a"

ref_b_fn_prefix="FSC/ref_b"

ptcl_a_fn_prefix="subtomograms/subtomo"

ptcl_b_fn_prefix="subtomograms/subtomo"

weight_a_fn_prefix="otherinputs/ampspec"

weight_b_fn_prefix="otherinputs/ampspec"

weight_sum_a_fn_prefix="FSC/wei_a"

weight_sum_b_fn_prefix="FSC/wei_b"

output_fn_prefix="FSC/ref_auto_b"

tomo_row="7"

iclass="0"

fsc_mask_fn="FSC/fsc_mask.em"

filter_a_fn=""

filter_b_fn=""

pixelsize=1

nfold=1

rand_threshold=0.8

plot_fsc=1

do_sharpen=1

box_gaussian=1

filter_mode=1

filter_threshold=0.143

plot_sharpen=1

do_reweight=0