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First, configuration (or simply config) files must be ASCII text files. On Windows systems, these files typically end with the .txt extension, but any file name or extension is acceptable. You can design and edit your config files with any simple text editor, such as Notepad or Wordpad in Windows, or vi, vim, nano, pico, and numerous other tools on GNU/Linux. Typically all config files should start with two important lines:
-mdat \your_path\ISOTOPE.DAT
-hdat \your_path\Hardklor.dat
These lines are paths to the data files required by Hardklör for operation. This is particularly important
if you store Hardklör in a central location (such as /usr/bin/ or C:\Program Files\), but wish to operate it
on mass spectrometry files stored in a different folder. In the above example, simply replace "your_path" with the
path of your ISOTOPE.DAT and Hardklor.dat files.
Also essential to every config file is a line or several lines of execution. In their basic form, lines of execution need only an input file and an output file:
MyData1.ms1 MyResults1.txt
MyData2.ms1 MyResults2.txt
In the example above, two files will be analyzed: MyData1.ms1 and MyData2.ms1. MyData1.ms1 will be analyzed first
and the results will be stored in a tab-delimited text file called MyResults1.txt. After analysis is finished on
this first file, analysis will start again on MyData2.ms1 and will be stored in MyResults2.txt.
To optimize your analysis, it will be necessary to set parameters for your data analysis. Parameters are represented in two ways: Global and Local. Global parameters are applied to every data file being analyzed. Local parameters are applied to only one data file. You can mix your parameters so that some are applied globally, but others are applied locally. Global parameters are represented as lines starting with a dash. You may put only one global parameter per line in your config file:
-corr 0.90
-d 3
The lines above set two parameters globally. The first parameter sets the correlation threshold to 0.90. The
second parameter sets a maximum deconvolution (depth) to 3 overlapping peptides. These two parameters will be
applied to every line of execution that comes after them.
Local parameters are set on the same line as the lines of execution. You can have as many parameters on a single line as necessary in these cases:
MyData1.ms1 MyResults1.txt -corr 0.90 -d 3
MyData2.ms1 MyResults2.txt -corr 0.95 -d 3
In the example above, two lines of execution are shown to have different parameters. MyData1.ms1 will be analyzed
with a correlation threshold of 0.90 and a maximum depth of 3. MyData2.ms1 will be analyzed with a correlation
threshold of 0.95 and a maximum depth of 3.
Notice in the last example that both MyData1.ms1 and MyData2.ms1 are analyzed with a maximum depth of 3. It would therefore be more efficient to set this parameter globally, but keep the correlation threshold parameters local. This can be done as shown in the following example:
-d 3
MyData1.ms1 MyResults1.txt -corr 0.90
MyData2.ms1 MyResults2.txt -corr 0.95
In order for the maximum depth of 3 to be applied to both data files, it must be set before the lines of
execution.
Now that you have a basic understanding of how a config file works, you might want to know what the most commonly used parameters are:
-a
-cdm
-chMin
-chMax
-corr
-d
-p
-res
-sn
-snWin
-win
For optimal Hardklör performance,
you should always set the above parameters to values appropriate for your data.
Details for each of these parameters can be found in the documentation on this website. These are the most
important parameters because they 1) Characterize your data files, 2) Specify what you want found in your
data files, and 3) Set the basic thresholds for finding isotope distributions. Typically, these parameters are set
globally when analyzing several data files from the same instrument.
For the fastest performance, two parameters should always be set as such:
-a FastFewestPeptides
-cdm Q
-a FastFewestPeptides specifies Hardklör to return the fewest overlapping peptides to fit a distribution, and
to perform the analysis using as much memory as possible to maximize speed. This parameter should be changed only
if the system has low memory. -cdm Q tells Hardklör to use the QuickCharge algorithm to find charge
state. This is much faster than other computation methods such as a Fast-Fourier Transform.
Additionally, increasing the values of the -chMax, -d, or -p parameters increases computation time, so it is recommended not to set them higher than necessary for your analysis. Decreasing the value of -snWin also increases computation time.
There are two ways to run Hardklör from the command-line. The first is to specify the data file and parameters you wish to use. The second is to specify a configuration file that contains parameters and a list of one or more files to analyze. The configuration file is the recommended method for two reasons: 1) It is simpler to set the large number of parameters this way, and 2) It allows you to batch multiple analyses without user interaction. To execute Hardklör, simply type the following command from your command prompt:
Hardklor -conf your_config.txt
Where your_config.txt is the name of your configuration file. Please see the section on
Sample Config Files or the config file setup tutorial
for instructions on how to make a configuration file.
Krönik must be run from the command-line. It is available for both Windows and GNU/Linux. For a quick help guide,
you can type Additionally, you can adjust some parameters for Krönik. Parameters must be set on the command line prior to your Hardklör results file. There are currently four parameters as follows:
A persistent PID is defined as a set of PIDs that: A detailed README file is included in the download of Krönik.
Hardklör is Copyright ©2007 University of Washington. All rights reserved. Written by Michael R. Hoopmann, Michael J. MacCoss, in the Department of Genome Sciences at the University of Washington. |
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