Best way to view wig/bigwig/bedgraph files - because IGV failing...

[KUMAR Sujai]

Apologies for cross posting (not sure if the ABC mailing list is just for the journal club or for random questions too - list managers please advise! :-))

I have a bam file with TOO many reads (25M) mapped to a 236 kb viral genome - as you can imagine IGV falls over while trying to load as BAM - presumably because the depth is too much (?) at each position...

So I tried converting bam to bigwig/bedgraph, and the files look right. but when I try to load those in IGV - nothing actually shows up in the coverage area.

Has this happened to anyone else?

If so, what do you use to show depth of coverage of reads mapped across a small genome (or small bit of a genome) - another tool? Or a custom python/R/perl script? If so, would you mind sharing it? (I could make the plot in anything, but it takes longer to get it to look pretty than to make the plot so am hoping to cheat and use someone else's  :-))

Thanks!

- Sujai

The University of Edinburgh is a charitable body, registered in Scotland, with registration number SC005336.

[Kashyap Chhatbar]

Hi Sujai,

What tool are you using for converting to bigwig? I would recommend this https://deeptools.readthedocs.io/en/develop/content/tools/bamCoverage.html

Also, make sure you put effective genome size https://deeptools.readthedocs.io/en/latest/content/feature/effectiveGenomeSize.html according to the viral genome.

This tool creates flawless bigwigs (at least for mammalian genomes).

Unless, Of course you would have tried this already, ignore this.

Best,

Kashyap

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[CEZARD Timothe]

Have not used it but this might be useful

https://github.com/brentp/goleft/tree/master/indexcov

 

Cheers,

Tim

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From: <ashworth-c...@googlegroups.com> on behalf of KUMAR Sujai <sujai...@ed.ac.uk>
Reply-To: "ashworth-c...@googlegroups.com" <ashworth-c...@googlegroups.com>
Date: Wednesday, 24 April 2019 at 16:54
To: Ashworth Codemonkeys <ashworth-c...@googlegroups.com>, "ashworth-bioin...@googlegroups.com" <ashworth-bioin...@googlegroups.com>, "bla...@googlegroups.com" <bla...@googlegroups.com>
Subject: [ashworth-code-monkeys] Best way to view wig/bigwig/bedgraph files - because IGV failing...

 

Apologies for cross posting (not sure if the ABC mailing list is just for the journal club or for random questions too - list managers please advise! :-))

 

I have a bam file with TOO many reads (25M) mapped to a 236 kb viral genome - as you can imagine IGV falls over while trying to load as BAM - presumably because the depth is too much (?) at each position...

 

So I tried converting bam to bigwig/bedgraph, and the files look right. but when I try to load those in IGV - nothing actually shows up in the coverage area.

 

Has this happened to anyone else?

 

If so, what do you use to show depth of coverage of reads mapped across a small genome (or small bit of a genome) - another tool? Or a custom python/R/perl script? If so, would you mind sharing it? (I could make the plot in anything, but it takes longer to get it to look pretty than to make the plot so am hoping to cheat and use someone else's  :-))

 

Thanks!

 

- Sujai

The University of Edinburgh is a charitable body, registered in Scotland, with registration number SC005336.

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[Dominik R. Laetsch]

On 24 Apr 2019, at 16:53, KUMAR Sujai <sujai...@ed.ac.uk> wrote:

Apologies for cross posting (not sure if the ABC mailing list is just for the journal club or for random questions too - list managers please advise! :-))

I'll allow it ... 😉

I have a bam file with TOO many reads (25M) mapped to a 236 kb viral genome - as you can imagine IGV falls over while trying to load as BAM - presumably because the depth is too much (?) at each position...

So I tried converting bam to bigwig/bedgraph, and the files look right. but when I try to load those in IGV - nothing actually shows up in the coverage area.

Well, I like old-school samtools tview for looking at things...

Has this happened to anyone else?

If so, what do you use to show depth of coverage of reads mapped across a small genome (or small bit of a genome) - another tool? Or a custom python/R/perl script? If so, would you mind sharing it? (I could make the plot in anything, but it takes longer to get it to look pretty than to make the plot so am hoping to cheat and use someone else's  :-))

Do you want a genome browser or do you want to compute coverages? For the latter I think indexcov or https://github.com/brentp/mosdepth are the best options, mosdepth does cumulative coverage decay plots ...

Cheers, 

Dom

Thanks!

- Sujai

The University of Edinburgh is a charitable body, registered in Scotland, with registration number SC005336.

[Dominik R. Laetsch]

Re mosdepth: if you go for this, you can use the output and modify the attached python script to make a cumulative-cov-decay-plot (see PNG) from the output ...

[KUMAR Sujai]

Many thanks to all. Will try all suggestions and report back (yes, I needed to visualise it, not just calculate depth, which bedtools did fine too)

Thanks Dom/Verity for confirming appropriate use - so if anyone reading this is on codemonkeys but not on ABC, it might be worth signing up to ABC (ashworth-bioinforma...@googlegroups.com or visit this group at https://groups.google.com/group/ashworth-bioinformatics-club)

On Wed, 24 Apr 2019 at 11:22, Dominik R. Laetsch <dominik...@gmail.com> wrote:

Re mosdepth: if you go for this, you can use the output and modify the attached python script to make a cumulative-cov-decay-plot (see PNG) from the output ...

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On 24 Apr 2019, at 17:07, 'Verity Hill' via Ashworth Bioinformatics Club <ashworth-bioin...@googlegroups.com> wrote:

Hi Sujai,

We envisioned the ABC mailing list to be for the journal club but also as a forum to answer questions just like this! We thought it could draw together code monkeys and bioinformatics journal club

happy questioning

(I do not know the answer to your question)

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Sujai Kumar

The University of Edinburgh

Ashworth Laboratories

Charlotte Auerbach Road

Edinburgh EH9 3FL, UK

[Edward Wallace]

I've started using R/tidyverse for this.

library(tidyverse)

library(valr)

plot_bgmribo <- function(bg,chromr,startr,endr) {

        ggplot(data=bg %>%

                   filter(chrom==chromr, start >= startr, end <= endr),

               aes(ymin=0,ymax=value,xmin=start,xmax=end)) +

        geom_rect(fill="blue",colour=NA) +

        labs(y="read count", x="position on chromosome")

}

bg <- read_bedgraph("mybed.bedgraph")

Can also get .gff files in R from rmonad vignette https://cran.r-project.org/web/packages/rmonad/vignettes/gff-processing.html

And plot them analogously, let me know if you want a script.

I haven't tried with bigwig.

Best

Edward

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