cnt file - library size

[Magda Maslon]

I would like to use scale factors (obtained from spike - ins quantification) to adjust my profiles in ngs plot. You suggest using .cnt files to this effect. When are these files created? I cannot locate these files, I am also not sure how to use them as an input?

[Clayton Collings]

Hi,

The .cnt files are created after you run it one time.  These files are ~7-12 bytes in size.

The .cnt files contain the number of mapped reads that meet some default quality score.

You can adjust the number of reads in this file and rerun ngsplot.  The new heatmaps

and metaplots RPMs will then be adjusted.  For example, if you find that the .cnt file

indicates that the sample has 10M read, and the spike-in indicates that the signal of the

sample should be decreased by 50%, then manually change .cnt file so that it has 20M reads

instead of 10M.  After you save this substitution, run ngsplot again.  Usually the .cnt files

are located in the working directory.  See powerpoint presentation I made about chipseq

spike-in normalization when I presented a paper about it at a lab meeting 5 years ago if

you are confused about calculations.  Hope this helps.

Best,

Clayton

On Fri, Jul 22, 2022 at 2:51 AM Magda Maslon <magdam...@gmail.com> wrote:

I would like to use scale factors (obtained from spike - ins quantification) to adjust my profiles in ngs plot. You suggest using .cnt files to this effect. When are these files created? I cannot locate these files, I am also not sure how to use them as an input?

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Clayton K. Collings, PhD

Computational Biologist

Kadoch Laboratory

Department of Pediatric Oncology

Dana-Farber Cancer Institute

Harvard Medical School

Broad Institute of MIT and Harvard

[Magda Maslon]

Thank you for your response.

This is helpful, especially re .cnt file localization - I was trying to find that file where the output was, but it is indeed stored in the working directory.

The ppt is very helpful, but I did get confused with your explanation. In your example: the spike-in indicates that the signal should be decreased by 50%, in that case should we not change from 10M to 5M, rather than to 20M?

thank you for your help

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