Import Peptide Error

support
Import Peptide Error rbao1357  2018-02-06 16:23
 
Hi guys,

I am a new user of Skyline (and to the MS field). I have been experiencing an error over the last 2 weeks when attempting to import peptides in the initial steps.

I have previously created technical replicate files under names (RB1, RB1b, RB1c etc), under the (.group), (wiff), (.wiff.scan) formats. These files are all stored in a single folder. When attempting to extract chromatograms, the peptide search is duplicating the first (RB1.wiff) file, rather than finding (RB1b etc) files. I have tested this with a number of different files, and the software is not locating the other files under different file names. I have also tested against (RB1) and (RB2) only, and it successfully builds the peptide spectrum.

This generates the below error.

An item with the same key has already been added.

Skyline version: 4.1.0.11714 (64-bit)
Installation ID: cf11b43b-5d9e-48e1-b3b6-ba6bcb35d288
Exception type: ArgumentException
Error message: An item with the same key has already been added.

--------------------

Stack trace:

   at System.ThrowHelper.ThrowArgumentException(ExceptionResource resource)
   at System.Collections.Generic.Dictionary`2.Insert(TKey key, TValue value, Boolean add)
   at pwiz.Skyline.Model.Results.MeasuredResults.set_Chromatograms(IList`1 value) in c:\proj\skyline_4_1_x64\pwiz_tools\Skyline\Model\Results\MeasuredResults.cs:line 80
   at pwiz.Skyline.Model.Results.MeasuredResults..ctor(IList`1 chromatograms, Boolean disableJoining) in c:\proj\skyline_4_1_x64\pwiz_tools\Skyline\Model\Results\MeasuredResults.cs:line 57
   at pwiz.Skyline.SkylineWindow.ImportResults(SrmDocument doc, List`1 namedResults, String optimize) in c:\proj\skyline_4_1_x64\pwiz_tools\Skyline\SkylineFiles.cs:line 2601
   at pwiz.Skyline.SkylineWindow.ModifyDocumentInner(Func`2 act, Action onModifying, Action onModified) in c:\proj\skyline_4_1_x64\pwiz_tools\Skyline\Skyline.cs:line 671
   at pwiz.Skyline.SkylineWindow.ModifyDocument(String description, IUndoState undoState, Func`2 act, Action onModifying, Action onModified) in c:\proj\skyline_4_1_x64\pwiz_tools\Skyline\Skyline.cs:line 635
   at pwiz.Skyline.SkylineWindow.ModifyDocument(String description, Func`2 act) in c:\proj\skyline_4_1_x64\pwiz_tools\Skyline\Skyline.cs:line 629
   at pwiz.Skyline.FileUI.PeptideSearch.ImportPeptideSearchDlg.WizardFinish() in c:\proj\skyline_4_1_x64\pwiz_tools\Skyline\FileUI\PeptideSearch\ImportPeptideSearchDlg.cs:line 606
   at pwiz.Skyline.FileUI.PeptideSearch.ImportPeptideSearchDlg.NextPage() in c:\proj\skyline_4_1_x64\pwiz_tools\Skyline\FileUI\PeptideSearch\ImportPeptideSearchDlg.cs:line 358
   at System.Windows.Forms.Control.OnClick(EventArgs e)
   at System.Windows.Forms.Button.OnClick(EventArgs e)
   at System.Windows.Forms.Button.OnMouseUp(MouseEventArgs mevent)
   at System.Windows.Forms.Control.WmMouseUp(Message& m, MouseButtons button, Int32 clicks)
   at System.Windows.Forms.Control.WndProc(Message& m)
   at System.Windows.Forms.ButtonBase.WndProc(Message& m)
   at System.Windows.Forms.Button.WndProc(Message& m)
   at System.Windows.Forms.NativeWindow.Callback(IntPtr hWnd, Int32 msg, IntPtr wparam, IntPtr lparam)

Exception caught at:
   at System.Windows.Forms.Application.ThreadContext.OnThreadException(Exception t)
   at System.Windows.Forms.Control.WndProcException(Exception e)
   at System.Windows.Forms.NativeWindow.Callback(IntPtr hWnd, Int32 msg, IntPtr wparam, IntPtr lparam)
   at System.Windows.Forms.UnsafeNativeMethods.DispatchMessageW(MSG& msg)
   at System.Windows.Forms.UnsafeNativeMethods.DispatchMessageW(MSG& msg)
   at System.Windows.Forms.Application.ComponentManager.System.Windows.Forms.UnsafeNativeMethods.IMsoComponentManager.FPushMessageLoop(IntPtr dwComponentID, Int32 reason, Int32 pvLoopData)
   at System.Windows.Forms.Application.ThreadContext.RunMessageLoopInner(Int32 reason, ApplicationContext context)
   at System.Windows.Forms.Application.ThreadContext.RunMessageLoop(Int32 reason, ApplicationContext context)
   at System.Windows.Forms.Form.ShowDialog(IWin32Window owner)
   at pwiz.Skyline.SkylineWindow.ShowImportPeptideSearchDlg(Nullable`1 workflowType) in c:\proj\skyline_4_1_x64\pwiz_tools\Skyline\SkylineFiles.cs:line 2866
   at pwiz.Skyline.SkylineWindow.importPeptideSearchMenuItem_Click(Object sender, EventArgs e) in c:\proj\skyline_4_1_x64\pwiz_tools\Skyline\SkylineFiles.cs:line 2853
   at System.Windows.Forms.ToolStripItem.RaiseEvent(Object key, EventArgs e)
   at System.Windows.Forms.ToolStripMenuItem.OnClick(EventArgs e)
   at System.Windows.Forms.ToolStripItem.HandleClick(EventArgs e)
   at System.Windows.Forms.ToolStripItem.HandleMouseUp(MouseEventArgs e)
   at System.Windows.Forms.ToolStrip.OnMouseUp(MouseEventArgs mea)
   at System.Windows.Forms.ToolStripDropDown.OnMouseUp(MouseEventArgs mea)
   at System.Windows.Forms.Control.WmMouseUp(Message& m, MouseButtons button, Int32 clicks)
   at System.Windows.Forms.Control.WndProc(Message& m)
   at System.Windows.Forms.ToolStrip.WndProc(Message& m)
   at System.Windows.Forms.ToolStripDropDown.WndProc(Message& m)
   at System.Windows.Forms.NativeWindow.Callback(IntPtr hWnd, Int32 msg, IntPtr wparam, IntPtr lparam)
   at System.Windows.Forms.UnsafeNativeMethods.DispatchMessageW(MSG& msg)
   at System.Windows.Forms.UnsafeNativeMethods.DispatchMessageW(MSG& msg)
   at System.Windows.Forms.Application.ComponentManager.System.Windows.Forms.UnsafeNativeMethods.IMsoComponentManager.FPushMessageLoop(IntPtr dwComponentID, Int32 reason, Int32 pvLoopData)
   at System.Windows.Forms.Application.ThreadContext.RunMessageLoopInner(Int32 reason, ApplicationContext context)
   at System.Windows.Forms.Application.ThreadContext.RunMessageLoop(Int32 reason, ApplicationContext context)
   at pwiz.Skyline.Program.Main(String[] args) in c:\proj\skyline_4_1_x64\pwiz_tools\Skyline\Program.cs:line 311


I have attached screenshots of the way the Skyline is behaving in a pptx. Are you able to provide any insight?

Thanks,

Rebecca
 
 
Nick Shulman responded:  2018-02-06 16:46
The error is happening because Skyline is trying to give two different replicates the same name.

Usually Skyline is smart enough to append a number to a replicate name if it would conflict with an existing replicate, but it looks like that feature is not working correctly for you.

I am not sure whether that has something to do with the particular names of the samples inside of your multi-sample wiff files, or whether that "do you want to remove the common prefix and suffix" dialog is messing things up.

It would be very helpful if you could send us your .wiff, .wiff.scan files and your Skyline document.

In Skyline, you can use the menu item:
File > Share > (complete)
to create a .zip file containing your Skyline document and supporting files.

You can attach that .zip file, as well as your .wiff and .wiff.scan files to this support request. Alternatively, you can upload those files here: https://skyline.ms/files.url

Thank you so much for contacting us. We saw the error reports coming in but we had no way to contact you to figure out how to reproduce this error.
 
rbao1357 responded:  2018-02-06 17:54
Hi Nick,

I have attached my Skyline document that I created from files (RB1) and (RB2). Due to the error from (RB1) and (RB1b), a document could not be created. I have attached the (.wiff) and (.wiff.scan) files in a separate Zip.

Thanks,

Rebecca
 
rbao1357 responded:  2018-02-06 18:41
 
rbao1357 responded:  2018-02-06 22:59
Hi Nick,

Files were too large to attach to this support request, so I have uploaded them to the shared folder under names (RBAO.sky.zip) and (RBAO ZIP.rar).

Thanks,

Rebecca
 
Nick Shulman responded:  2018-02-07 17:00
Thank you for sending me those files.

I believe Skyline is having trouble because some of your .wiff files had the same name as each other at the time that your .group files were created.
That is, you file "rb1_result.group" came from a file called "...20171025\RB1.wiff" and "rb1b_result.group" came from "...20171026\RB1.wiff".
When Skyline tries to match the .group files up with the .wiff files that you have, Skyline thinks both of them are associated with "RB1.wiff" that you have, and Skyline ignores the "RB1b.wiff" that you now have.

Skyline appears to have a bug in it where it then tries to create two replicates called "RB1", which causes the error that you see.

The good news is that you can made it almost all of the way through the Import Peptide Search wizard, and you can just ignore the error. The only step that you have left to do is extracting chromatograms, which you can do with the Skyline menu item:
File > Import > Results

The bad news is that Skyline is not going to incorrectly associate the peptide spectrum matches from "rb1b_result.group" with the file "rb1.wiff", and Skyline will not believe that there were any peptide spectrum matches in "rb1b.wiff". Because of this, Skyline's peak picking will not be as good as it should be.