I am trying to covert my bam file into bedgraph using bedtools. For that I sorted the bam file by names, and then run the follwoing commands:
bedtools bamtobed -bedpe -i IgG_H3K4m3.sortedByName.bam > IgG_H3K4m3.bed
awk '$1==$4 && $6-$2 < 1000 {print $0}' IgG_H3K4m3.bed > IgG_H3K4m3.clean.bed
cut -f 1,2,6 IgG_H3K4m3.clean.bed | sort -k1,1 -k2,2n -k3,3n > IgG_H3K4m3.frag.bed
bedtools genomecov -bg -i IgG_H3K4m3.frag.bed -g Mmu.GrCm38.chromSize > IgG_H3K4m3.bedgraph
But with the last command I get the following error
Input error: Chromosome 12 found in non-sequential lines. This suggests that the input file is not sorted correctly.
When I look what the command before last is doing ( the sorting steps) I can see a strange behavior of sort :
cut -f 1,2,6 P193/Mmu.GrCm38/bowtie2/IgG_H3K4m3.clean.bed | sort -k1,1 -k2,2n -k3,3n| cut -f 1 | uniq
10
1
11
12
1
14
15
16
1
18
2
3
5
6
9
MT
X
Somehow it can't sort chromosome 1 correctly. Is there any wat to explain this ( or even better to correct it).
thanks
•
written
by
Assa Yeroslaviz
Hey Assa.
You could try this, and it should work just fine:
sort -V -k1,1 -k2,2n -k3,3n
Example:
cat test.bed
1 12 32
2 12 76
12 12 765
12 12 54
10 453 987
11 1 34
11 34 88
MT 34 100
MT 22 2222
X 12222 5555665
12 3324 77566
Y 676 89898989
sort -k1,1 -k2,2n -k3,3n test.bed
1 12 32
10 453 987
11 1 34
11 34 88
12 12 54
12 12 765
12 3324 77566
2 12 76
MT 22 2222
MT 34 100
X 12222 5555665
Y 676 89898989
sort -V -k1,1 -k2,2n -k3,3n test.bed
1 12 32
2 12 76
10 453 987
11 1 34
11 34 88
12 12 54
12 12 765
12 3324 77566
MT 22 2222
MT 34 100
X 12222 5555665
Y 676 89898989
Kevin
Thanks this seems to work, but is there an explanation as to why the other one didn't work int his case?
I cannot confirm but it's something to do with the version of sort that you have.
It is odd, I have the same version as Assa Yeroslaviz (on Mac) and cannot reproduce this behaviour.
I'm working on an ubuntu server with version 16.0.4. Maybe there is a difference. But with the -V option it does what it suppose to do. thanks again
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Output of
sort --version?sort (GNU coreutils) 8.22