View source for Routine 2D Experiment

&nbsp;'''NMR Data Processing &gt; Routine 2D Experiments via NMRPipe '''<br> 

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===== '''Brief Description'''  =====

NMRPipe is designed for multidimensional NMR processing.&nbsp; The preferred method of processing data is to compile the commands and functions into a script file.&nbsp; The script file can be easily modified for different type of experiments and serves as a record of how the data was processed.&nbsp; An example of converting the 2D HSQC spectrometer data to NMRPipe format is shown along with processing and visualizing of the dataset. 

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===== '''Software Information'''  =====

NMRPipe (download information and user manual) 

<span style="text-decoration: underline">[http://%22 http://spin.niddk.nih.gov/NMRPipe/]</span> 

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Brief descriptions of specific functions are accessible via nmrPipe – 

‘nmrPipe –help’ will list most functions. 

‘nmrPipe –fn GM –help’ will give description of the GM function. 

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Supported Platforms 

Linux (RedHat Linux/Fedora)&nbsp; 

Mac OS X (10.3.4 and up) 

SGI Irix&nbsp; (6.2 and up) 

Sparc Solaris (2 and up) 

Windows XP Pro with Microsoft Services for UNIX (SFU 3.5). 

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===== '''Converting Spectrometer Data into NMRPipe Format'''  =====

To convert data from spectrometer format to NMRPipe format is usually done using the build-in program called ‘varian’ for a Varian dataset ('bruker' for a Bruker dataset and 'delta' for dataset from JEOL). &nbsp; [[Brief_description_of_philosophy,_commands,_and_functions_of_NMRPipe|Brief summaries of the build-in programs can be found here.]]

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Step 1:&nbsp; Under the same directory where the procpar and fid files are, type ‘varian’ and this will start a tcl/tk script that brings up two windows. 

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[[Image:Varian-utility-raw.png]] [[Image:Varian-script-raw.png]] 

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Step 2: Click on the ‘Read Parameters’ button and the script will read the parameter file (‘procpar’) and update the parameters. &nbsp; 

<br> [[Image:Varian-utility-readed.png]] [[Image:Varian-script-readed.png]] <br> 

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The template shows parameters for two dimensions x and y.&nbsp; The x dimension is the direct dimension and is always collected as complex data (quadrature detection) and the xMODE is set as complex.&nbsp; The xN (real+imaginary points) parameter is twice the xT (complex points) parameter.&nbsp; For the indirect dimension y, the yN parameter is twice the yT parameter (number of increments, ni for a Varian instrument)<span style="font: 12.0px Times New Roman">,</span> indicating that we are collecting complex dataset to account for ‘quadrature’ in the indirect dimension.&nbsp; For the sensitivity-enhanced experiments the yMODE will be set to “Rance-Kay”.&nbsp; The frequency values can be modified using the pull down tab.&nbsp; The center position ppm of each dimension (xCAR and yCAR) can be updated at this stage and it can also be update later. 

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Step 3: Click on ‘Save Script’, and the ‘Execute Script’.&nbsp; A converted dataset in NMRPipe format will be created as test.fid along with an UNIX shell script called ‘fid.com’.&nbsp; All of the processing parameters and the location of the input and output files are store in the ‘fid.com’ script and it can be modified using a text editor.&nbsp; &nbsp; 

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===== '''Processing and Visualizing 2D Dataset'''  =====

Similar to the conversion script, all of the commands for data processing are also complied into a script file (brief summaries of the commands).&nbsp; The nmrDraw program has a macro-editing tool to aid in the construction of script.&nbsp; The nmrDraw program can be launch by typing ‘nmrDraw &amp;’ under the same directory the dataset is.&nbsp; The macro editor can be started by selecting ‘Macro Edit’ from the ‘File’ menu.&nbsp; This editor contains build in macros to process 2D, 3D, and 4D dataset<span style="font: 12.0px Times New Roman">,</span> or it can create lines of text for various functions.&nbsp; The script can be saved and executed inside the macro editor. &nbsp; 

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[[Image:NMRPipe-Macro-Editors.png]] 

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An alternative way to create these processing script is to copy an existing one and edit it with a text editor.&nbsp; An example of processing script for a 2D 15N HSQC spectrum is shown below.&nbsp; The script starts off by read in the time domain data ‘test.fid’ using the ‘nmrPipe –in’ command following by ‘\’ at the end of each line.&nbsp; Each subsequent ‘nmrPipe –fn’ command line will begin with a unix ‘|’ and end with ‘\’. &nbsp; A solvent suppression filter SOL was first applied to removed residual solvent followed by a 90 degrees sine bell square apodization (equivalent to cosine) with scale factor for first point as 0.5.&nbsp; The data is then zero<span style="font: 12.0px Times New Roman">-</span>filled to the nearest power of 2, Fourier transformed, phased, and the amide region of the spectrum (left half) extracted.&nbsp; The TP functions transpose the data to the y-axis to apply linear prediction, apodization, zero fill, Fourier transform, phasing to the y-dimension, and output the finished file as ‘test.ft2’.&nbsp; When the first point is collected at half dwell time (for Varian instrument, f1180=’y’), the phasing values in the indirect dimension is set to ‘-90 for p0 and 180 for p1’ and the first point multiplier set to 1.0<span style="font: 12.0px Times New Roman">.</span> 

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#!/bin/csh

nmrPipe&nbsp; -in test.fid &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; \ 

| nmrPipe -fn SOL &nbsp; \ ## Solvent suppresion 

| nmrPipe&nbsp; -fn SP -off 0.5 -end 1.00 -pow 2 -c 0.5&nbsp; \ ## Apodization 

| nmrPipe&nbsp; -fn ZF -auto &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; \ ## Zero fill 

| nmrPipe&nbsp; -fn FT -auto &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; \ ## Fourier transformation 

| nmrPipe -fn PS -p0 13.6 -p1 0.0&nbsp; -di&nbsp; &nbsp; \ ## Phase correction 

| nmrPipe&nbsp; -fn EXT –left –sw &nbsp; \ # Extract spectrum 

| nmrPipe -fn TP&nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; \ ## Transpose to the y-axis 

| nmrPipe -fn LP -fb &nbsp; &nbsp; &nbsp; &nbsp; \ ## Add liner prediction 

| nmrPipe -fn SP -off 0.5 -end 0.98 -pow 2 -c 1.0&nbsp; \ 

| nmrPipe -fn ZF -auto &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; \ 

| nmrPipe -fn FT -auto&nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; &nbsp; \ 

| nmrPipe -fn PS -p0 -90.0 -p1 180.0 &nbsp; -di &nbsp; &nbsp; \ 

| nmrPipe -fn TP&nbsp; &nbsp; &nbsp; &nbsp; \ 

| nmrPipe –out test.ft2&nbsp; -verb 2 -ov 

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===== Suggested Workflow  =====

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Step 1: Generate the processing script with the appropriate commands and functions using the macro editor or modified existing script.&nbsp; Set both the p0 and p1 phasing values of the direct dimension to 0. 

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Step 2: Execute the processing script in a UNIX terminal.&nbsp; Load the spectrum into nmrDraw to perform phase correction and optimizing the applied functions.&nbsp; A 1D horizontal trace can be activate by typing ‘h’ in the spectrum window and the phase of the dimension can be adjusting using the P0 and P1 slider bars.&nbsp; 
[[Image:NMRDraw-Phasing.png]]

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Step 3: Execute the processing script with optimized parameters. 

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Step 4: Convert the processed data to the format of graphical NMR assignment program. 
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