goleft_linux64 covstats error: panic: EOF about goleft

[brentp]

can you re-index the bam and try again?

[YiweiNiu]

Thank you for your quick reply.

I tried to re-create the bam index using samtools index, but found EOF marker is absent. So, I re-generated the bam/index. It's solved now.

[javaidm]

Hi,
I've installed the 'goleft' from source, and trying to run the 'indexcov' with the following command (it's index is present in the same dir):
goleft indexcov --directory indexcov my_bam.bam

But I'm getting the error below:

`panic: sam: duplicate field: line 2: "@pg\tID:bwa\tPN:bwa\tVN:0.7.15-r1140\tCL:bwa mem -t 32 -R @rg\tID:TZ5637P\tSM:TZ5637P\tPL:ILLUMINA\tPU:TZ5637P\tLB:TZ5637P /Shares/layer_shared/ref/hg38/GRCh38_full_analysis_set_plus_decoy_hla.fa TZ5637P_TD180603257_HT3LKCCXY_L3_1.fq.gz TZ5637P_TD180603257_HT3LKCCXY_L3_2.fq.gz"

goroutine 1 [running]:
github.com/brentp/goleft/indexcov.RefsFromBam(0x7fff13dff9cf, 0x5b, 0x0, 0x0, 0xc0002660d0, 0x7fff13dff9c6, 0x8)
/Users/maja5467/go/src/github.com/brentp/goleft/indexcov/indexcov.go:321 +0x4c4
github.com/brentp/goleft/indexcov.getReferences(0x7fff13dff9c6, 0x8, 0xc00022ae01)
/Users/maja5467/go/src/github.com/brentp/goleft/indexcov/indexcov.go:338 +0xbe
github.com/brentp/goleft/indexcov.Main()
/Users/maja5467/go/src/github.com/brentp/goleft/indexcov/indexcov.go:403 +0x1e3
main.main()
/Users/maja5467/go/src/github.com/brentp/goleft/cmd/goleft/goleft.go:68 +0x170
`
Any thoughts?

Regards,
Mahmood Javaid.

[brentp]

this error is from biogo/hts, I think it's finding ID:bwa and ID:TZ... and thinking that's a duplicate tag. probably because you used \t instead of the expanded, literal tab when you called bwa
maybe @kortschak could verify?

the solution is to re-header or change the way you call bwa mem.

[javaidm]

Thank you for your prompt reply, for the time being, I'll probably re-run the bwa-mem.

[kortschak]

@javaidm Can you post the complete panic output in a <pre> block (triple back tick block), and paste the first few lines of the SAM header also in a code block.

It looks to me like you have a corrupt header with an @pg and @rg line on the same line. Also, I'm a little confused that these are lower case; the SAM spec requires that these be upper case.

Oh, I see what the problem is, the -R flag takes "@rg\tID:TZ5637P\tSM:TZ5637P\tPL:ILLUMINA\tPU:TZ5637P\tLB:TZ5637P" as a param. This is not correctly being interpreted. I don't see a sensible way to deal with this kind of horror. Looking at the bwa manual, the string is literally quoted, are you doing this? From what I can see it looks like you should do

bwa mem -t 32 -R '@rg\tID:TZ5637P\tSM:TZ5637P\tPL:ILLUMINA\tPU:TZ5637P\tLB:TZ5637P' /Shares/layer_shared/ref/hg38/GRCh38_full_analysis_set_plus_decoy_hla.fa

but you seem to be doing

bwa mem -t 32 -R "@rg\tID:TZ5637P\tSM:TZ5637P\tPL:ILLUMINA\tPU:TZ5637P\tLB:TZ5637P" /Shares/layer_shared/ref/hg38/GRCh38_full_analysis_set_plus_decoy_hla.fa

Can you confirm?

[javaidm]

panic: sam: duplicate field: line 2: "@PG\tID:bwa\tPN:bwa\tVN:0.7.15-r1140\tCL:bwa mem -t 32 -R @RG\tID:TZ5637P\tSM:TZ5637P\tPL:ILLUMINA\tPU:TZ5637P\tLB:TZ5637P /Shares/layer_shared/ref/hg38/GRCh38_full_analysis_set_plus_decoy_hla.fa TZ5637P_TD180603257_HT3LKCCXY_L3_1.fq.gz TZ5637P_TD180603257_HT3LKCCXY_L3_2.fq.gz"

goroutine 1 [running]:
github.com/brentp/goleft/indexcov.RefsFromBam(0x7ffd8e65f690, 0x5b, 0x0, 0x0, 0xc0001e4340, 0x7ffd8e65f689, 0x6)
	/Users/maja5467/go/src/github.com/brentp/goleft/indexcov/indexcov.go:321 +0x4c4
github.com/brentp/goleft/indexcov.getReferences(0x7ffd8e65f689, 0x6, 0xc0001aad01)
	/Users/maja5467/go/src/github.com/brentp/goleft/indexcov/indexcov.go:338 +0xbe
github.com/brentp/goleft/indexcov.Main()
	/Users/maja5467/go/src/github.com/brentp/goleft/indexcov/indexcov.go:403 +0x1e3
main.main()
	/Users/maja5467/go/src/github.com/brentp/goleft/cmd/goleft/goleft.go:68 +0x170```

I can confirm that the string is not being literally quoted at the moment (it's part of a Nextflow pipeline, so I need to find out a way to both literally quote it and achieve the variable interpolation). Anyhow, for now I managed to run ```goleft indexcov``` on cram index, but will look into running the 'bwa-mem' as prescribed in their manual. Thanks for your help!