running ASAPratio for SILAC data

[sudarshan kumar]

Can anyone please help me know, why when I am running express and ASAPratio it always takes more than two days time and though express is completed but after running for around 80% in asapratio it stucks and shows only the running status but nothing moves ahead.  

[David Shteynberg]

Dear Sudarshan,

It is hard to say why this particular execution resulted in apparent failure, but if you are willing to compress the whole directory of this analysis and send me a download link, I would be happy to download this dataset and check why it failed.

Cheers!

-David

On Wed, Nov 22, 2023 at 12:04 PM sudarshan kumar <kumarsu...@gmail.com> wrote:

Can anyone please help me know, why when I am running express and ASAPratio it always takes more than two days time and though express is completed but after running for around 80% in asapratio it stucks and shows only the running status but nothing moves ahead.  

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[sudarshan kumar]

Thank you so much David,

But can you tell me how can i compress? Do you mean this link?

(6) ISB/SPC Trans Proteomic Pipeline :: jobs

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The real voyage of discovery consists not in seeking new lands but seeing with new eyes. — Marcel Proust

Dr. Sudarshan Kumar

(Fulbright-Nehru Fellow)
(B.V.Sc.& A.H., M.V.Sc., PhD.)
Sr. Scientist
Animal Biotechnology Center
(Proteomics and Cell Biology Lab.)
National Dairy Research Institute Karnal, 132001
Haryana, India
Contact No 09254912456
URL www.ndri.res.in

Orcid Id: https://orcid.org/0000-0002-9816-4307

[sudarshan kumar]

EXECUTING: cd c:/TPP/data/SILAC/Comet_SILAC/Comet_silac_param && c: && C:/TPP/bin/xinteract -Ninteract_silaccomtwofiles.pep.xml -p0.0 -l6 -THREADS=1 -PPM -OA -ipP -X-m0.1-c5-p1-L-nR,10.008269-nK,8.014199 -A-lRK-F-C-Z-r0.05 110623_Silac_TEST_01.pep.xml 110623_Sud_LH_TEST_02.pep.xml

[David Shteynberg]

Hello Sudarshan,

Please zip this directory on your system: c:/TPP/data/SILAC/Comet_SILAC/Comet_silac_param

Then upload the zip file to your favorite cloud server and share the link with me.

Thanks!

-David

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[sudarshan kumar]

I have sent it through google drive

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[sudarshan kumar]

Hi David,

could you get time to see into my analysis? I need your advice.

Best regards,

Sud

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[David Shteynberg]

Dear Sud,

I was able to process your dataset!  Although, I am still working on getting a faster version of ASAPRatio compiled for your testing purposes.  Meanwhile you can help the process by filtering your files so that zero intensity peaks in the MS1 data are removed and the MS1 data is centroided.  You can accomplish both of these things using the msconvert tool.  This will help reduce the time requirement to process this dataset.  I should have more to say about this particular case in the next few days.

Cheers!

-David

On Nov 29, 2023, at 10:17 AM, sudarshan kumar <kumarsu...@gmail.com> wrote:

Hi David,

could you get time to see into my analysis? I need your advice.

Best regards,

Sud

On Wed, Nov 22, 2023 at 3:51 PM 'David Shteynberg' via spctools-discuss <spctools...@googlegroups.com> wrote:

Hello Sudarshan,

Please zip this directory on your system: c:/TPP/data/SILAC/Comet_SILAC/Comet_silac_param

Then upload the zip file to your favorite cloud server and share the link with me.

Thanks!

-David

On Wed, Nov 22, 2023 at 3:17 PM sudarshan kumar <kumarsu...@gmail.com> wrote:

EXECUTING: cd c:/TPP/data/SILAC/Comet_SILAC/Comet_silac_param && c: && C:/TPP/bin/xinteract -Ninteract_silaccomtwofiles.pep.xml -p0.0 -l6 -THREADS=1 -PPM -OA -ipP -X-m0.1-c5-p1-L-nR,10.008269-nK,8.014199 -A-lRK-F-C-Z-r0.05 110623_Silac_TEST_01.pep.xml 110623_Sud_LH_TEST_02.pep.xml

On Wed, Nov 22, 2023 at 3:07 PM sudarshan kumar <kumarsu...@gmail.com> wrote:

Thank you so much David,

But can you tell me how can i compress? Do you mean this link?

(6) ISB/SPC Trans Proteomic Pipeline :: jobs

On Wed, Nov 22, 2023 at 2:31 PM 'David Shteynberg' via spctools-discuss <spctools...@googlegroups.com> wrote:

Dear Sudarshan,

It is hard to say why this particular execution resulted in apparent failure, but if you are willing to compress the whole directory of this analysis and send me a download link, I would be happy to download this dataset and check why it failed.

Cheers!

-David

On Wed, Nov 22, 2023 at 12:04 PM sudarshan kumar <kumarsu...@gmail.com> wrote:

Can anyone please help me know, why when I am running express and ASAPratio it always takes more than two days time and though express is completed but after running for around 80% in asapratio it stucks and shows only the running status but nothing moves ahead.  <Screenshot 2023-11-22 120242.png>

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[sudarshan kumar]

Thank you so much . I will try the way have suggested.

Best regards

Sud

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[sudarshan kumar]

can i select all the three options?

Will it impact Express analysis?

Centroid all scans (MS1 and MS2) -- meaningful only if data was acquired in profile mode
Compress peak lists for smaller output file
Write the output as a gzipped file (other TPP tools can read gzipped files directly)

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[sudarshan kumar]

Hi David,

I am using this tutorial for Expressan dASAPratio . can yuo please comment on if I am using the right one.?

Regards,

Sud 

[sudarshan kumar]

one more query-

I used same samples - for label free and SILAC express?

Ideally I should get a similar conclusion from both the experiments.

But the differentially expressed proteins are different in the two analyses.

NOte: I have used only one run file of control and one run file of treatment for LFQ

While for SILAC I have used 6 run files of same samples (technical replicates).

Please let me know where I am wrong? In my point of view increasing the number of run files will improve PSM and hence quantification using express. What should i believe my SILAC express results or the same sample LFQ (st. peters results)?

 

[David Shteynberg]

Hopefully, I am understanding your queries correctly, here goes.  Centroiding the data will reduce the number of peaks in the spectra and reduce the running time of the analysis.  Compressing the files to make them smaller will require that the tools reading the files uncompress the data (usually internally) prior to using it which will increase the running time.  As far as affecting the results, compressing the files should not change the data while centroiding will. As far as how much this will affect the final ratios probably depends on the quality of centroiding algorithm and the quality of the data.  Run some tests yourself and get a feel on some data you know well;  testing should help you refine your approach.  Hope that helps!

-David

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[David Shteynberg]

Hello Sudarshan, again!

I was able to update the ASAPRatioPeptideParser.exe tool for your testing purposes to see if it runs a bit faster after some optimization I was able to implement.  Please download the following version: https://drive.google.com/file/d/1Pe1pBFZ9BOxrrYNY6O8rkrybKaGtDhZZ/view?usp=sharing

And replace the copy you have in C:/TPP/bin/ 

When I ran it on my not very powerful desktop computer it completed on your data in about 42,000 seconds using the new code (less than 12 hours)

Notice that the interface still says that "This command is still running..." even though the log says "job completed" 

This is because for very long jobs the webserver times-out before the job completes.  To solve this the user must click the link just above that is next to the text: "If you commands have actually completed but the server timed out, click here"

I hope this helps you process your data given your current computational resources in less than 1 day.

Cheers!

-David

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[sudarshan kumar]

Thank you so much David for doing this for me. In fact I was running asapratio again on the centroid mzml file. But I see that though express completed easily but the asapratio is still running. Is there any way in tpp to see how much time will it take to complete. 

But I will try with the version you have shared the link with me. Should I use centroid mzml for this ?.

Regards, Sudarshan 

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[David Shteynberg]

Of course, I am glad to help.  The old and the new versions should run faster on the centroided data.  There is no way to tell a priori with ASAPRatio how long it will take, but since it ran to completion on my machine and it generates one dot per ten PSMs it has processed; based on the dots you can see how much of the ASAPRatio processing has completed.  Here are the dots from my run on your data:

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