Target region coverage

[shmibble]

Hi,
I am performing an alignment and am interested in several target regions. I am interested in finding out the minimum and maximum coverage in each region (meaning the base with the minimum coverage and the base with the maximum coverage in each region) for quality control purposes. I already have a sorted bam file.
Does anyone know how I can do this? Is there a program that can help?

Thanks!

[amitm]

hi,
Using bedtools coverageBed with the -hist option would give you base level coverage. (contiguous bases with same depth would be collapsed in one entry though)

http://bedtools.readthedocs.org/en/latest/content/tools/coverage.html

[shmibble]

Thanks for the reply.
I don't really understand what I need file B for. How do I get a histogram of the coverage of file A (a sorted bam file)?

Thanks again.

[amitm]

hi,
You would need two things -
1) Post-alignment, coord. sorted BAM
2) BED file of your target regions

Then for each of your target region (or sub-parts of it if there are different depth areas in a given target) you would get the number of reads (which is depth or coverage) aligned.

A BED file at its simples is a tab-delimited 3 col. file having Chr. Start & End coords

[shmibble]

Hi,

Thanks for the reply and sorry it took me a while to respond.
It still doesn't seem to work for me... Here's the command I type:

bedtools coverage -abam -a aligned_sorted.bam -b ref.bed

It says ERROR: Unrecognized Parameter: aligned_sorted.bam

If I remove the -abam option I get a different error: "Unexpected file format. Please use tab-delimited BED, GFF or VCf. Perhaps you have non-integer starts or ends at line 1?"
I'm not really sure what this means. My bed file is a 3 column tab delimited file. Each line looks like "chr10 123456789 234567891".

Any advice?

Thanks again

[amitm]

hi,
Quickly -
The -abam and -a parameter are mutually xclusive. Use one.
If you have two BED files, say, and you want to calculate coverage of one over other. Then its like
-a First.bed -b Second.bed

Else if you have a BAM file and a BED file, then
-abam Input.bam -b Sorted.bed

Hope that clears it.

My first guess for the 2nd error is that that the chromosome name format in your BAM and BED don't match.
Check using
samtools view -H Input.bam