Question: GATK BaseRecalibrator No overlapping contigs found ERROR
0
gravatar for marongiu.luigi
20 months ago by
Germany, Mannheim, UMM
marongiu.luigi520 wrote:

Dear all,

I have extracted reads that did not mapped to a reference genome with

samtools view -h -f 4 <aln-srt>.bam <unmap-srt>.bam

where the input file is sorted. Then I created the fastq files with:

bamToFastq -i <unmap-srt>.bam -fq <Ump_1>.fq -fq2 <Ump_2>.fq
gzip <Ump_1>.fq
gzip <Ump_2>.fq

This created many warning errors such as:

*****WARNING: Query HWI-ST1437:64:C3UM1ACXX:4:2315:4948:54750 is marked as paired, but its mate does not occur next to it in your BAM file.  Skipping.

but overall, the procedure was completed and I could re-align to another reference with:

bwa mem -t 8 -R $RG <ref.fa> <Ump_1>.fq <Ump_2>.fq | \
samtools sort -o <newaln-srt>.bam

and I got alignments such as this (from IGV): enter image description here

So far so good, i'd say (unless there is a flaw in this process of course).

I then tried to apply a recalibration with the following steps:

sambamba markdup -r --overflow-list-size 1000000 --hash-table-size 1000000 \
<newaln-srt>.bam <newaln-srt-dedup>.bam
gatk BaseRecalibrator \
  -R <ref>.fa \
  -I <newaln-srt-dedup>.bam \
  -O <newaln-srt-dedup>.tab \
  --known-sites <ref>.vcf.gz

But I got:

[...]
Running:
[...]
.BaseRecalibrator done. Elapsed time: 0.51 minutes.
Runtime.totalMemory()=1161822208
***********************************************************************

A USER ERROR has occurred: Input files reference and reads have incompatible contigs: No overlapping contigs found.
  reference contigs = [1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, X, Y, MT, CHR_HSCHR15_4_CTG8, CHR_HSCHR6_MHC_SSTO_CTG1, CHR_HSCHR6_MHC_MCF_CTG1, [...]
423.1, KI270392.1, KI270394.1]
  reads contigs = [V]

***********************************************************************
Set the system property GATK_STACKTRACE_ON_USER_EXCEPTION (--java-options '-DGATK_STACKTRACE_ON_USER_EXCEPTION=true') to print the stack trace.

So the question is: how can I deal with this error? Shall I do the recalibration in the first place?

Thank you.

ADD COMMENTlink modified 20 months ago by Pierre Lindenbaum129k • written 20 months ago by marongiu.luigi520
2
gravatar for Pierre Lindenbaum
20 months ago by
France/Nantes/Institut du Thorax - INSERM UMR1087
Pierre Lindenbaum129k wrote:

reads contigs = [V]

your read is mapped on a contig named "V'

USER ERROR has occurred: Input files reference and reads have incompatible contigs: No overlapping contigs found. reference contigs = [1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, X, Y, MT, CHR_HSCHR15_4_CTG8, CHR_HSCHR6_MHC_SSTO_CTG1, CHR_HSCHR6_MHC_MCF_CTG1, [...]

but the reference doesn't contain such contig named V

how can I deal with this error?

use the rererence that was used to map with bwa. If it is the same, then there is a problem with your bam. show use the header+first reads of the bam please.

ADD COMMENTlink written 20 months ago by Pierre Lindenbaum129k

Good spot, it must be something with the headers in the reference. I'll try again. Thank you

ADD REPLYlink written 20 months ago by marongiu.luigi520
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