GATK BaseRecalibrator - Input files knownSites and reference have incompatible contigs

Question

[cl10101]

I am trying to use GATK BaseRecalibrator but I get this error: "Input files knownSites and reference have incompatible contigs"

##### ERROR MESSAGE: Input files knownSites and reference have incompatible contigs. Please see https://www.broadinstitute.org/gatk/guide/article?id=63for more information. Error details: The contig order in knownSites and reference is not the same; to fix this please see: (https://www.broadinstitute.org/gatk/guide/article?id=1328),  which describes reordering contigs in BAM and VCF files..

##### ERROR   knownSites contigs = [chr1, chr10, chr10_GL383545v1_alt, chr10_GL383546v1_alt, chr10_KI270824v1_alt, chr10_KI270825v1_alt, chr11, chr11_GL383547v1_alt, chr11_JH159136v1_alt, chr11_JH159137v1_alt, ......

##### ERROR   reference contigs = [chr1, chr10, chr11, chr11_KI270721v1_random, chr12, chr13, chr14, chr14_GL000009v2_random, chr14_GL000225v1_random, chr14_KI270722v1_random, chr14_GL000194v1_random, .....

I searched the Internet and found out that I should use the compatible vcf file and reference, but I downloaded vcf file from hg38 bundle ftp://ftp.broadinstitute.org/bundle/hg38/hg38bundle/ so I suppose it should be compatible. My reference uses 'chr1,etc.', while the vcf file uses '1,2,etc.', so I added chr to the vcf file. I also used Picard ReorderSam on my BAM file and SortVcf on vcf file as described here http://gatkforums.broadinstitute.org/gatk/discussion/1328/errors-about-contigs-in-bam-or-vcf-files-not-being-properly-ordered-or-sorted. Now it looks like my reference is sorted differently (lexicographically sorted). I have no idea how can I fix this problem, should I sort my reference or download other reference file?

Answer 1

[Pierre Lindenbaum]

don't change your bam.

you only need to convert the names in your vcf files

• VCF files: Change Chromosome Notation
• Changing Chromosome Notation On Vcf
• https://github.com/lindenb/jvarkit/wiki/VcfRenameChromosomes

and the use picard SortVcf using the REF= fasta.dict file as the reference for sorting.

[cl10101]

Thank you, but I already used this option running SortVcf: java -Xmx3g -jar ../picard.jar SortVcf I=dbsnp_144.hg38_with_chr.vcf O=dbsnp_144.hg38_with_chr_sorted.vcf SEQUENCE_DICTIONARY=../ref/hg38.dict and the problem still exists.

[Pierre Lindenbaum]

can you please show me the output of the following commands:

$ grep "##contig" -m 10 dbsnp_144.hg38_with_chr.vcf 
$ grep "##contig" -m 10 ../ref/hg38.dict 
$ grep "##contig" -m 10  dbsnp_144.hg38_with_chr_sorted.vcf

[cl10101]

$ grep "##contig" -m 10 dbsnp_144.hg38_with_chr.vcf

$ grep "##contig" -m 10 ../ref/hg38.dict

$ grep "##contig" -m 10 dbsnp_144.hg38_with_chr_sorted.vcf

##contig=<ID=chr1,length=248956422>

##contig=<ID=chr10,length=133797422>

##contig=<ID=chr11,length=135086622>

##contig=<ID=chr11_KI270721v1_random,length=100316>

##contig=<ID=chr12,length=133275309>

##contig=<ID=chr13,length=114364328>

##contig=<ID=chr14,length=107043718>

##contig=<ID=chr14_GL000009v2_random,length=201709>

##contig=<ID=chr14_GL000225v1_random,length=211173>

##contig=<ID=chr14_KI270722v1_random,length=194050>

[Pierre Lindenbaum]

and head ../ref/hg38.dict ?

[cl10101]

@HD     VN:1.5  SO:unsorted
@SQ     SN:chr1 LN:248956422    M5:2648ae1bacce4ec4b6cf337dcae37816     UR:file:/home/user/ref/hg38.fa
@SQ     SN:chr10        LN:133797422    M5:907112d17fcb73bcab1ed1c72b97ce68     UR:file:/home/user/ref/hg38.fa
@SQ     SN:chr11        LN:135086622    M5:1511375dc2dd1b633af8cf439ae90cec     UR:file:/home/user/ref/hg38.fa
@SQ     SN:chr11_KI270721v1_random      LN:100316       M5:9654b5d3f36845bb9d19a6dbd15d2f22     UR:file:/home/user/ref/hg38.fa
@SQ     SN:chr12        LN:133275309    M5:e81e16d3f44337034695a29b97708fce     UR:file:/home/user/ref/hg38.fa
@SQ     SN:chr13        LN:114364328    M5:17dab79b963ccd8e7377cef59a54fe1c     UR:file:/home/user/ref/hg38.fa
@SQ     SN:chr14        LN:107043718    M5:acbd9552c059d9b403e75ed26c1ce5bc     UR:file:/home/user/ref/hg38.fa
@SQ     SN:chr14_GL000009v2_random      LN:201709       M5:862f555045546733591ff7ab15bcecbe     UR:file:/home/user/ref/hg38.fa
@SQ     SN:chr14_GL000225v1_random      LN:211173       M5:63945c3e6962f28ffd469719a747e73c     UR:file:/home/user/ref/hg38.fa

Answer 2

[erwan.scaon]

I stumbled across something similar recently.

I finally managed to get rid of the "incompatible contigs" error : COSMIC vcf file compatibility for Mutect2