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Variant Discovery in High-Throughput Sequencing Data

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Developed in the Data Sciences Platform at the Broad Institute, the toolkit offers a wide variety of tools with a primary focus on variant discovery and genotyping. Its powerful processing engine and high-performance computing features make it capable of taking on projects of any size. Learn more

Receiving zero variants processed after HaplotypeCaller and Select Variants

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    Genevieve Brandt (she/her)

    Hi Candace Grimes,

    It looks like you do have some sites in your gvcf that are variant sites (Not reference blocks):

    ptg000002l 368020 . A G,<NON_REF> 65.83 . DP=3;ExcessHet=3.0103;MLEAC=1,0;MLEAF=0.500,0.00;RAW_MQandDP=2937,3 GT:AD:DP:GQ:PL:SB 1/1:0,3,0:3:9:79,9,0,79,9,79:0,0,1,2

    As of now, I don't think there are issues with the HaplotypeCaller step.

    Have you run GenotypeGVCFs yet? I would recommend running that tool and then seeing the output to determine if your variant sites are expected. I'm thinking that your SelectVariants commands didn't pick up the variant sites because you restricted the command to only SNPs and INDELs, and you haven't genotyped the gvcf yet.

    Let me know what you find.

    Best,

    Genevieve

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    Candace Grimes

    Yes, thank you Genevieve! I believe that worked. I am now getting variants processed.

    I do have another question: I am trying to analyze shorter sequences with this same genome. They have a low mapping rate (~10%), and when I run them through the HaplotypeCaller tool, I receive the USER ERROR: Input files reference and reads have incompatible contigs: No overlapping contigs found. I checked the duplication metrics and the percent duplication seems to be above 80%. 

    ## METRICS CLASS picard.sam.DuplicationMetrics
    LIBRARY UNPAIRED_READS_EXAMINED READ_PAIRS_EXAMINED SECONDARY_OR_SUPPLEMENTARY_RDS UNMAPPED_READS UNPAIRED_READ_DUPLICATES READ_PAIR_DUPLICATES READ_PAIR_OPTICAL_DUPLICATES PERCENT_DUPLICATION ESTIMATED_LIBRARY_SIZE
    Unknown Library 73608 0 0 544979 64370 0 0 0.874497

    Please let me know if you have any suggestions and thank you again! 

    All the best, 

    Candace

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    Candace Grimes

    Thank you Genevieve! 

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