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Variant called if HC is run on individual bam but variant not called on multiple input bam
I encounter a situation when ;
If I run Haplotype caller on one .bam file - Haplotype caller called the mutation (sanger validated)
java GenomeAnalysisTK.jar -T HaplotypeCaller -nct 15 -minPruning 2 \ -R genome.fa \ -I sample_1.bam \ -L TP53.bed \ -U ALLOW_N_CIGAR_READS \ -o sample_1.vcf
But, If i run Haplotype caller with multiple bam as input - Haplotype caller failed to call the mutation in the sample mentioned above
java GenomeAnalysisTK.jar -T HaplotypeCaller -nct 15 -minPruning 2 \ -R genome.fa \ -I sample_1.bam -I sample_2.bam -I sample_3 .bam ..... -I sample_19.bam \ -L TP53.bed \ -U ALLOW_N_CIGAR_READS \ -o multisample.vcf
I used the -bamOutput option to view the .bam file HC would expect to see when calling the mutation. Please find below IGV screenshot
java GenomeAnalysisTK.jar -T HaplotypeCaller -nct 15 -minPruning 2 \ -R genome.fa \ -I sample_1.bam -I sample_2.bam -I sample_3 .bam ..... -I sample_19.bam \ -L TP53.bed \ -U ALLOW_N_CIGAR_READS \ --bamOutput multiple_all.bam --bamWriterType ALL_POSSIBLE_HAPLOTYPES \ -o multisample.vcf
1st row is HC run only on sample_1.bam
2nd row is HC run on multiple.bam input - using the --bamOutput option alone
3rd row is HC run on multiple.bam input - using the --bamOutput option + --bamWriterType ALL_POSSIBLE_HAPLOTYPES
The coverage on the 1st row (IGV) of sample1 is 262 (Ref(C):3, Mut(A):259)
The coverage on the 3rd row (IGV) file at the position of interest is 3627(Ref(C):3411, Mut(A):215, G:1 , DEL:13)
Could the reason why HC unable to call the mutation (when multiple.bam are used as input) is because the postion of interest is has too much noise ~3 base upstream? (shown in screen shot below)
Any help understanding why mulisample bam input would fail to call the variant would me most helpful.
Thanks in advance