Aaron: /* Merging all integrated images */

2015-08-07T17:15:38Z

Merging all integrated images

← Older revision		Revision as of 17:15, 7 August 2015
Line 125:		Line 125:
	significance_filter.apply = True		significance_filter.apply = True
	set_average_unit_cell = True		set_average_unit_cell = True
			pixel_size = 0.11
	mysql {		mysql {
	database = <i>db_name</i>		database = <i>db_name</i>
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	$ cxi.xmerge L498-merge.phil		$ cxi.xmerge L498-merge.phil
	Merging statistics are printed on standard output. The merged MTZ-file is written to a file whose name is determined by the value of <code>output.prefix</code> in the configuration file (with the values shown above, the output file would be <code>L498_thermolysin.mtz</code>). Note that the version of merging programs from 28 March, 2013 do <em>not</em> not report the <i>R</i><sub>split</sub> statistic.		Merging statistics are printed on standard output. The merged MTZ-file is written to a file whose name is determined by the value of <code>output.prefix</code> in the configuration file (with the values shown above, the output file would be <code>L498_thermolysin.mtz</code>). Note that the version of merging programs from 28 March, 2013 do <em>not</em> not report the <i>R</i><sub>split</sub> statistic.


	== References ==		== References ==

	<references/>		<references/>

Aaron: /* Indexing the secondary lattice */

2014-04-19T12:26:10Z

Indexing the secondary lattice

← Older revision		Revision as of 12:26, 19 April 2014
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	=== Indexing the secondary lattice ===		=== Indexing the secondary lattice ===

	Indexing the secondary lattice is very similar to indexing the primary lattice, but requires a change to the source code. Edit <code><b><i>/path/to</i></b>/phenix-src-20130328/labelit_regression/xfel/xfel_targets.py</code>, and uncomment (<i>i.e.</i> remove the leading <code>#</code> character) <code>"~~outlier_detection_switch~~=True"</code> on line 25. Then edit the configuration file, <code><i>L498-indexigrate.cfg</i></code> above, and change <i>integration-first-lattice</i> to <i>integration-second-lattice</i> in order not to overwrite the results of the previous analysis of the primary lattice. Before reanalyzing the first batch, ensure that <code>xtal_target</code> is set to <code>thermolysin27</code>.		Indexing the secondary lattice is very similar to indexing the primary lattice, but requires a change to the source code. Edit <code><b><i>/path/to</i></b>/phenix-src-20130328/labelit_regression/xfel/xfel_targets.py</code>, and uncomment (<i>i.e.</i> remove the leading <code>#</code> character) <code>"outlier_detection.switch=True"</code> on line 25. Then edit the configuration file, <code><i>L498-indexigrate.cfg</i></code> above, and change <i>integration-first-lattice</i> to <i>integration-second-lattice</i> in order not to overwrite the results of the previous analysis of the primary lattice. Before reanalyzing the first batch, ensure that <code>xtal_target</code> is set to <code>thermolysin27</code>.
	$ cxi.pyana -c <i>L498-indexigrate.cfg</i> <b><i>/path/to/xtc/files</i></b>/e157-r0016-*.xtc		$ cxi.pyana -c <i>L498-indexigrate.cfg</i> <b><i>/path/to/xtc/files</i></b>/e157-r0016-*.xtc
	$ cxi.pyana -c <i>L498-indexigrate.cfg</i> <b><i>/path/to/xtc/files</i></b>/e157-r0017-*.xtc		$ cxi.pyana -c <i>L498-indexigrate.cfg</i> <b><i>/path/to/xtc/files</i></b>/e157-r0017-*.xtc
Line 109:		Line 109:
	$ cxi.pyana -c <i>L498-indexigrate.cfg</i> <b><i>/path/to/xtc/files</i></b>/e157-r0073-*.xtc		$ cxi.pyana -c <i>L498-indexigrate.cfg</i> <b><i>/path/to/xtc/files</i></b>/e157-r0073-*.xtc
	The number of integrated secondary lattices should be close to 2,021, the number reported in the ''cctbx.xfel'' paper<ref name="Hattne:2014"/>.		The number of integrated secondary lattices should be close to 2,021, the number reported in the ''cctbx.xfel'' paper<ref name="Hattne:2014"/>.


	== Merging all integrated images ==		== Merging all integrated images ==

Hattne: Cleanups, spell checks, etc.

2014-03-14T18:09:00Z

Cleanups, spell checks, etc.

Show changes

Hattne: Created instructions to reproduce L498 thermolysin processing results.

2014-03-10T10:54:28Z

Created instructions to reproduce L498 thermolysin processing results.

New page

This page contains instructions for reproducing the results reported in the 2014 ''cctbx.xfel'' paper<ref>[http://dx.doi.org/10.1038/nmeth.2887 Hattne, J et al. Accurate macromolecular structures using minimal measurements from X-ray free-electron lasers. Nat. Methods In press (2014).]</ref>. The data are available from the [http://cxidb.org/id-23 Coherent X-ray Imaging Data Bank (CXIDB)] (raw XTC files, 4.0 TiB), and must be downloaded to a local disk prior to starting the analysis. Furthermore, the PSDM Software Distribution must be [[Set up PSDM software | set up]], along with a test release and an empty analysis package. A [http://www.mysql.com MySQL] database is required to merge the integrated diffraction images. Processing time depends strongly on the computational resources available; using 48 processors on a 64-core 1.4 GHz Opteron-based computer, the analysis takes around 24 hours. The instructions assume some familiarity with ''cctbx.xfel'' and its installation and configuration procedure.

== Installing a ''cctbx.xfel'' snapshot from March 28, 2013 ==

The thermolysin data for the ''cctbx.xfel'' paper was processed around March 28, 2013. Unfortunately, regular nightly releases are not available for that time, but a specially prepared source-code bundle has been made available. This bundle differs from the regular ''cctbx'' bundles in that ''LABELIT'' is included, and that the directory layout is identical to that of a developer installation. To download and unpack the bundle in the current directory:
$ wget http://adder.lbl.gov/cctbx.xfel/downloads/cctbx.xfel-20130328.tar.gz
$ tar -xpvzf cctbx.xfel-20130328.tar.gz
Next, create a build directory (called <code>phenix-build-20130328</code> below, but that is an arbitrary choice), in which the sources are configured and compiled. The Python interpreter required to complete this step, must be the one supplied by the PSDM Software Distribution. Once the PSDM software has been [[Set up PSDM software | set up], this interpreter can be located using
$ find $SIT_ROOT/sw/external/python -perm /0111 -type f -wholename "*/$SIT_ARCH/*/python"
To prepare the build directory using this interpreter
$ mkdir phenix-build-20130328
$ cd phenix-build-20130328
$ python ../phenix-src-20130328/cctbx_project/libtbx/configure.py cxi_xdr_xes xfel
$ . setpaths.sh
where <code>python</code> is the path to the Python interpreter located using the previous find-command. Note that csh-users should run <code>source setpaths.csh</code> instead of <code>. setpaths.sh</code>. Then compile the sources
$ make
$ make
Note that the <code>make</code> command may need to be run twice in order to complete the build. Once <code>make</code> does not produce any output from the compiler, the build is complete.

To make the ''cctbx.xfel'' analysis modules available to PSDM's ''pyana'',
$ cd /path/to/test/release
$ sit_setup
$ cd my_ana_pkg
$ ln -fns /path/to/phenix-src-20130328/cctbx_project/xfel/cxi/cspad_ana src
$ cd ..
$ scons
where <code>/path/to/test/release</code> and <code>my_ana_pkg</code> are the path to the test release and the name of the analysis package chosen while [[Set up PSDM software | setting up the PSDM software distribution]]. <code>/path/to</code> denotes the path to the directory containing the unpacked ''cctbx.xfel'' sources. The last step compiles the ''cctbx.xfel'' analysis modules.

== Create a dark image ==

To meaningfully process the thermolysin diffraction data, an average of all the images in a dark run—a run without any X-rays impinging on the detector—must be subtracted from the individual diffraction images. The configuration file below can be used to produce such an average, as well as an image of the standard deviation of all the pixels over the course of the run.
# -*- mode: conf -*-

[pyana]
modules = my_ana_pkg.mod_average
num-cpu = 4

[my_ana_pkg.mod_average]
address = CxiDs1-0|Cspad-0
calib_dir = /path/to/phenix-src-20130328/cctbx_project/xfel/metrology/CSPad/run4/CxiDs1.0_Cspad.0
avg_basename = Ds1-avg
avg_dirname = r0031
stddev_basename = Ds1-stddev
stddev_dirname = r0031
/path/to refers to the directory containing the unpacked ''cctbx.xfel'' sources. All other options with values set in italics can be modified without adversely affecting averaging. The above file will use four simultaneous processes, and write the average and standard deviation images to files whose names start with <code>Ds1-avg</code> and <code>Ds1-stddev</code>, respectively, both in a directory called <code>r0031</code>.

The data deposited at the CXIDB contains a dark run, <code>r0031</code>. To average the images in that run, save the above configuration file to disk, e.g. <code>L498-dark.cfg</code>, apply modifications as necessary, and execute
$ cxi.pyana -c L498-dark.cfg /path/to/xtc/files/e157-r0031-*.xtc
where /path/to/xtc/files is the path to the directory containing the raw XTC files dowloaded from [http://cxidb.org/id-23 CXIDB]. The files written to the <code>r0031</code> directory will have a current datestamp appended, which can safely be removed to simplify subsequent configuration files.
$ cd r0031
$ mv Ds1-avg20140308104336073.pickle Ds1-avg.pickle
$ mv Ds1-stddev20140308104336371.pickle Ds1-stddev.pickle
$ cd ..
Further details are available on the [[Preparatory steps#Create a dark average|Create a dark image]] page of the [[Tutorials|tutorials]].

== Index the thermolysin data ==

A configuration file for processing the primary lattices in the thermolysin data is shown below.
# -*- mode: conf -*-

[pyana]
modules = my_ana_pkg.mod_hitfind:threshold \
my_ana_pkg.mod_hitfind:index
num-cpu = 48

[my_ana_pkg.mod_hitfind]
address = CxiDs1-0|Cspad-0
calib_dir = /path/to/phenix-src-20130328/cxi_xdr_xes/cftbx/metrology/CSPad/run4/CxiDs1.0:Cspad.0
dark_path = r0031/Ds1-avg.pickle
dark_stddev = r0031/Ds1-stddev.pickle
db_logging = False
detz_offset = 575

[my_ana_pkg.mod_hitfind:threshold]
dispatch = nop
distl_flags = permissive
distl_min_peaks = 16
threshold = 450
xtal_target = hitfind

[my_ana_pkg.mod_hitfind:index]
dispatch = index
integration_dirname = integration-first-lattice
integration_basename = int-
xtal_target = thermolysin27
The configuration file above instructs <code>mod_hitfind</code> to use 48 processes. It disables all image output, which reduces the amout of disk space required to perform the analysis to about 3.4 GiB. /path/to again refers to the directory containing the unpacked ''cctbx.xfel'' sources, and <code>dark_path</code> as well as <code>dark_stddev</code> may have to be changed to reflect the location of the previously generated dark images. Integration results will be written to the directory integration-first-lattice.

Due to particularities of the thermolysin measurement, processing proceeds in two batches. To analyse the first batch, runs 16 through 27, save the above configuration file to disk, e.g. <code>L498-indexigrate.cfg</code>, apply modifications as necessary, and execute
$ cxi.pyana -c L498-indexigrate.cfg /path/to/xtc/files/e157-r0016-*.xtc
$ cxi.pyana -c L498-indexigrate.cfg /path/to/xtc/files/e157-r0017-*.xtc
$ …
$ cxi.pyana -c L498-indexigrate.cfg /path/to/xtc/files/e157-r0027-*.xtc
where /path/to/xtc/files is the path to the directory containing the raw XTC files. The second batch, runs 71 through 73, was recorded using a different distance between the interaction region and the detector. This necessitates different areas of the detector to be ignored due to different shadowing, and this is accounted for by a different value of the <code>xtal_target</code> option in the configuration file. To analyse this set of runs, edit <code>L498-indexigrate.cfg</code>, change <code>thermolysin27</code> to <code>thermolysin73</code>, and
$ cxi.pyana -c L498-indexigrate.cfg /path/to/xtc/files/e157-r0071-*.xtc
$ cxi.pyana -c L498-indexigrate.cfg /path/to/xtc/files/e157-r0072-*.xtc
$ cxi.pyana -c L498-indexigrate.cfg /path/to/xtc/files/e157-r0073-*.xtc
On successful completion, the number of files in the integration-first-lattice directory corresponds to the number of successfully integrated images. Owing to variations in hardware and compiler internals, it may deviate slightly from 11,583, the number reported in the ''cctbx.xfel'' paper.

=== Index the secondary lattice ===

Indexing the secondary lattice is very similar to indexing the primary lattice, but requires a change to the source code. Edit <code>/path/to/phenix-src-20130328/labelit_regression/xfel/xfel_targets.py</code>, and uncomment (i.e. remove the leading <code>#</code> character) <code>"outlier_detection_switch=True"</code> on line 25. Then edit the configuration file, <code>L498-indexigrate.cfg</code> above, and change integration-first-lattice to integration-second-lattice in order not to overwrite the results of the previous analysis of the primary lattice. Before re-analysing the first batch, ensure that <code>xtal_target</code> is set to <code>thermolysin27</code>.
$ cxi.pyana -c L498-indexigrate.cfg /path/to/xtc/files/e157-r0016-*.xtc
$ cxi.pyana -c L498-indexigrate.cfg /path/to/xtc/files/e157-r0017-*.xtc
$ …
$ cxi.pyana -c L498-indexigrate.cfg /path/to/xtc/files/e157-r0027-*.xtc
Then set <code>xtal_target</code> is set to <code>thermolysin73</code> in the configuration file, and process the second batch.
$ cxi.pyana -c L498-indexigrate.cfg /path/to/xtc/files/e157-r0071-*.xtc
$ cxi.pyana -c L498-indexigrate.cfg /path/to/xtc/files/e157-r0072-*.xtc
$ cxi.pyana -c L498-indexigrate.cfg /path/to/xtc/files/e157-r0073-*.xtc
The number of integrated secondary lattices should be close to 2,021, the number reported in the ''cctbx.xfel'' paper.

== Merge all integrated images ==

The phil-file below defines values suitable for merging the primary and secondary lattices previously integrated.
data = integration-first-lattice
data = integration-second-lattice
d_min = 2.10
merge_anomalous = True
min_corr = -1
model = 2tli.pdb
nproc = 16
plot_single_index_histograms = False
raw_data.sdfac_auto = True
rescale_with_average_cell = True
significance_filter.apply = True
set_average_unit_cell = True
mysql {
database = db_name
passwd = db_passwd
runtag = L498_thermolysin
user = db_user
}
output {
n_bins = 10
prefix = L498_thermolysin
}
scaling {
algorithm = mark0
mtz_file = 2tli.mtz
show_plots = False
log_cutoff = 0.0
}
<code>integration-first-lattice</code> and <code>integration-second-lattice</code> may need to be adjusted to point to the directories where <code>mod_hitfind</code> left the integration results. <code>db_name</code><code>, db_user</code>, and <code>db_passwd</code> must be substituted with the database name and access credentials to a MySQL database. Databases on hosts other than the one used to merge the thermolysin data can be accessed by additionally specifing the <code>mysql.host</code> and <code>mysql.port</code> options. The model and structure factors for the scaling reference, <code>model</code> and <code>scaling.mtz_file</code> above, are both available for download from the [http://www.rcsb.org RCSB Protein Data Bank (PDB)]. If the [http://www.phenix-online.org ''PHENIX''] suite is installed, this is conveniently achieved from the command line using
$ phenix.fetch_pdb --mtz 2tli

To merge the thermolysin data, save the suitably modified configuration to e.g. <code>L498-merge.phil</code>, and run
$ cxi.merge L498-merge.phil
$ cxi.xmerge L498-merge.phil
Merging statistics are printed on standard output. The merged MTZ-file is written to a file whose name is determined by the value of <code>output.prefix</code> in the configuration file (<code>L498_thermolysin.mtz</code> with the values shown above). Note that the version of merging programs from 28 March, 2013 do not not report the Rsplit statistic.

== References ==

<references/>

Processing L498 thermolysin - Revision history

Aaron: /* Merging all integrated images */

Aaron: /* Indexing the secondary lattice */

Hattne: Cleanups, spell checks, etc.

Hattne: Created instructions to reproduce L498 thermolysin processing results.