HIFI READS PROVIDE A COMPREHENSIVE VIEW OF VARIATION IN THE HUMAN GENOME 5 Mb 3 Mb 10 Mb 1 bp SNVs ≥50 bp structural variants (SVs) 1-49 bp indels HiFi reads Short reads vs GRCh38
HIFI READS PROVIDE A COMPREHENSIVE VIEW OF VARIATION IN THE HUMAN GENOME 5 Mb 3 Mb 10 Mb 1 bp SNVs ≥50 bp structural variants (SVs) 1-49 bp indels HiFi reads Short reads vs GRCh38 SNVs and indels in difficult regions
HIFI READS PROVIDE A COMPREHENSIVE VIEW OF VARIATION IN THE HUMAN GENOME Short reads HiFi reads STRC STRC is a congenital deafness gene that requires long reads to cover all exons.
GOOGLE DEEPVARIANT IS A HIGHLY ACCURATE SMALL VARIANT CALLER FOR HIFI READS Poplin, R. E. et al. A universal SNP and small-indel variant caller using deep neural networks. Nat Biotechnol 25, 1097 (2018). -Variant calling pipeline powered by deep neural network -Fast and inexpensive -Run from binaries as well as Docker or Singularity containers -PacBio model trained on HiFi reads from Sequel and Sequel II Systems with median read quality >99.9% -Model is updated regularly to support PacBio Chemistry and Software updates
99.9 90 99 97 99.7 Accuracy, F1(%) precisionFDA Entries Top 12 entries and 25 of top 26 use PacBio HiFi reads Illumina Multi HiFi ONT HiFi DeepVariant Illumina DeepVariant Illumina GATK ONT DeepVariant
HIFI READS PROVIDE A COMPREHENSIVE VIEW OF VARIATION IN THE HUMAN GENOME 5 Mb 3 Mb 10 Mb 1 bp SNVs ≥50 bp structural variants (SVs) 1-49 bp indels HiFi reads Short reads vs GRCh38 Long indels and SVs genome-wide
CALL STRUCTURAL VARIANTS FROM HIFI READS WITH PBSV HiFi reads pbmm2 pbsv discover pbsv call variant calls (vcf) SMRT Link Structural Variant Calling SMRT Link Mapping OR
VARIANT DETECTION BENCHMARKING (HG002) Recall | Precision (%) HiFi Coverage SNVs Indels SVs 15-fold 99.53 | 99.89 95.16 | 96.23 97.41 | 94.48 30-fold 99.89 | 99.95 98.90 | 98.99 98.00 | 95.29 SNV and indel calls are from DeepVariant 1.0.0 and evaluated against the GIAB v4.2 small variant benchmark using Hap.py. SV calls are from pbsv 2.2.2 and evaluated against the GIAB v0.6 SV benchmark using Truvari.
PATHOGENIC VARIANTS DETECTED WITH HIFI READS Hiatt SM, Lawlor JMJ, et al. (2020). Long-read sequencing for the diagnosis of neurodevelopmental disorders. bioRxiv, doi:10.1101/2020.07.02.185447 Figure 1. Proband 6 has a de novo insertion resulting in duplication of exon 3 of CDKL5
COMPREHENSIVE VARIANT DETECTION WITH HIFI READS -HiFi = mappability of long reads + base quality of short reads -HiFi + DeepVariant yield most accurate small variant calls currently available with a single technology. -HiFi + pbsv yield highly accurate structural variant calls, including inversions, translocations, and copy number variants. -Recommend 15-fold coverage for most discovery applications. Datasets for the Ashkenazi trio (15 kb and 20 kb libraries) are deposited on SRA: HG002 (PRJNA586863) HG003 (PRJNA626365) HG004 (PRJNA626366)
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