how to use Indel religner against bam aligned to major allele human genome

[swNGS]

Hi,

I am looking at the difference in variant calls that I get having aligned my resequencing data to both the 1000 genomes reference genome human_g1k_v37.fasta and a major allle reference genome: Dewey et al, Plos Genetics.

For both standard and major reference genome alignments, I had planned to use GATK to call variants.
My question relates to whether I should be pointing GATK to either respective reference genome for each iteration, eg:

for major allele refernce alignment
java -Xmx4g -jar /path_to.../GenomeAnalysisTK/GenomeAnalysisTK.jar \
-I Major_allele_aligned_input_bam.bam \
-R /PATH_To_MAJOR_ALLELE_REF/CEUref.fasta \ <<----------- THIS
-T IndelRealigner \
-targetIntervals forIndelRealigner.intervals \
-o output_bam.bam \
--known /.../1000G_indels/1000G_biallelic.indels.b37.vcf \
--consensusDeterminationModel KNOWNS_ONLY \
-LOD 0.4

for 1000 genome ref alignment
java -Xmx4g -jar /path_to.../GenomeAnalysisTK/GenomeAnalysisTK.jar \
-I 1000_genomes_aligned_input_bam.bam \
-R /PATH_To_1000_GENOMES/human_g1k_v37.fasta \ <<----------- THIS
-T IndelRealigner \
-targetIntervals forIndelRealigner.intervals \
-o output_bam.bam \
--known /.../1000G_indels/1000G_biallelic.indels.b37.vcf \
--consensusDeterminationModel KNOWNS_ONLY \
-LOD 0.4

Thanks

[swNGS]

okay, that dosn't work!

GATK throws the following error, which I recollect getting in the past:


##### ERROR MESSAGE: Input files known and reference have incompatible contigs: No overlapping contigs found.
##### ERROR known contigs = [1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, X]
##### ERROR reference contigs = [chr1, chr2, chr3, chr4, chr5, chr6, chr7, chr8, chr9, chr10, chr11, chr12, chr13, chr14, chr15, chr16, chr17, chr18, chr19, chr20, chr21, chr22, chrX, chrY, chrM]

Its fairly obvious that the contigs are different, and I get something similar when I use the 1000 genomes reference:

##### ERROR MESSAGE: Input files reads and reference have incompatible contigs: No overlapping contigs found.
##### ERROR reads contigs = [chr1, chr2, chr3, chr4, chr5, chr6, chr7, chr8, chr9, chr10, chr11, chr12, chr13, chr14, chr15, chr16, chr17, chr18, chr19, chr20, chr21, chr22, chrX, chrY, chrM]
##### ERROR 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, GL000207.1, GL000226.1, GL000229.1, GL000231.1, GL000210.1, GL000239.1, GL000235.1, GL000201.1, GL000247.1, GL000245.1, GL000197.1, GL000203.1, GL000246.1, GL000249.1, GL000196.1,...etc

Does anyone have any idea how I can fix this ?

thanks

[aaronh]

Looks like the names of the chromosomes are not matching between the reference dictionary in the bam file, the reference fasta file and the known variants in the VCF file.

[swNGS]

Ah, that's helpful, I suspect it might be the known variants vcf file that's causing the problem. Would I have to edit the headers so that the contig names in both the major allele reference genome and the known variants in the VCF are the same ?

Also, and hopefully not a stupid question, what if one of these files refers to a contig that is missing in the other? I am thinking specifically of GL000207.1, GL000226.1... etc ?

Thanks,

Chris

[moty]

Hi there,
did you ever solve this? I am having the exact same problem.
I am using the hg19 reference fasta from the GATK website and got bam files as a given.

edit: I have tried renaming the chromosomes labels from the .dict and .fai files # --> chr# and ended up with a complaint the chrM is 2 read shorter.
If you have a wiser solution I'd love to hear it

Moty