What's wrong with my GATK HaplotypeCaller result without ALT

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Han Li

• January 02, 2022 14:39

Hello，I used the gatk 4.2.3.0 and STAR 2.7.9a, cellranger 6.1.2 to make the scRNA SNPs detection but i got a wrong. Here i used the bam from the cellranger 6.1.2 which use the star for mapping. And then, i used the haplotypeCaller for SNPs detection and here's the code and result.

/home/yph/anaconda3/envs/velocyto/bin/gatk --java-options "-Xmx20g" HaplotypeCaller \
--R /media/workspace/yph/cellranger-3.0.2/refdata-cellranger-GRCh38-3.0.0/fasta/genome.fa \
--I /media/workspace/yph/cellranger-3.0.2/A3_90/outs/possorted_genome_bam.bam \
--D /media/workspace/yph/cellranger-3.0.2/refdata-cellranger-GRCh38-3.0.0/resources_broad_hg38_v0_Homo_sapiens_assembly38.dbsnp138.rename.vcf \
--O /media/workspace/yph/LHY/A3_90.rename.g.vcf \
--ERC GVCF

/home/yph/anaconda3/envs/velocyto/bin/gatk --java-options "-Xmx20g" GenotypeGVCFs \
--R /media/workspace/yph/cellranger-3.0.2/refdata-cellranger-GRCh38-3.0.0/fasta/genome.fa \
--V /media/workspace/yph/LHY/A3_90.rename.g.vcf \
--O /media/workspace/yph/LHY/A3_90.rename.vcf

 

but in the result g.vcf file, the ALT column shows NON-REF and the position column shows something wrong? I had use the SplitNCigarReads for BAM file detection but nothing ERROR was found.  So ,i want to know why the haplotypeCaller could not find the SNPs information in the Cellranger-6.1.2a BAM file? Or , what can i do ?

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  Genevieve Brandt (she/her)

  • January 10, 2022 23:55

  Hi Han Li,

  Could you please share the example of the unexpected result? This will help to better identify what is going on.

  Thank you,

  Genevieve

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  王有栋

  • March 24, 2022 01:31

  Hello, I also have the above problem. In the GVCF file, ALT column is all "NON_REF", but in VCF, it is normal. Is there any reason for this

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  Genevieve Brandt (she/her)

  • April 06, 2022 21:19

  Hello!

  This looks like a normal GVCF file. NON_REF in the ALT column indicates that the position matches the reference. You are also not showing the same sites in your two screen shots, so that is why there is different info in the VCF and GVCF.

  You can read more about GVCFs and VCFs here:

  • https://gatk.broadinstitute.org/hc/en-us/articles/360035531812-GVCF-Genomic-Variant-Call-Format
  • https://gatk.broadinstitute.org/hc/en-us/articles/360035531692-VCF-Variant-Call-Format

  Let me know if you have any other questions.

  Best,

  Genevieve

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