Reveal true HLA allelic diversity
Characterizing polymorphisms in HLA genes outside of the protein binding regions is important for strengthening causality association studies1. High-sequence homology and hyper polymorphisms make it challenging to determine true HLA allelic diversity. Even though sequencing is becoming a more popular tool for HLA typing, diploid ambiguities and a reliance on imputation still persist. This is especially true when the length of homologous sequences between alleles extends beyond the capabilities of most sequencing platforms2.
Access the full spectrum of polymorphism across the entire HLA gene
Gain a deeper understanding of immune-related disease causality, graft-versus-host disease in hematopoietic transplantation, and drug hypersensitivity with Single Molecule, Real-Time (SMRT) Sequencing.
SMRT Sequencing delivers single-molecule observations with 10–15 kb reads capable of spanning the majority of HLA class I and II genes. Generate highly accurate consensus sequences (QV50 demonstrated) from amplicons spanning full-length HLA genes without a reference to obtain directly phased, high-resolution HLA types without imputation2.
Features of this application include the ability to:
- Flexibly scale project size (e.g., phase through either exons 2, 3 and/or 4, or the entire HLA gene) with cost-effective multiplexing solutions
- Phase polymorphisms with allele specificity across SNP-poor regions of HLA genes
- Achieve allele-level segregation without imputation
- Detect variants in regulatory regions within 5’ UTRs, introns, and 3’ UTRs
- Fully characterize minor variants in polyclonal samples, such as cancer or transcripts
- Obtain direct evidence for new HLA alleles through de novo, reference-free consensus generation
Workflow: from targeted region to accurate variant or haplotype
- Library preparation
- SMRT Sequencing on the PacBio Systems
- Data analysis with SMRT Analysis or PacBio DevNet
Featured research: Build complete gene references for HLA class II genes
Immunologists at DKMS generated reference-free consensus sequences of HLA-DPB 1 complete genes. At the time of the study, the IGMT/HLA database contained 12 full-length references out of 550 known HLA-DPB1 alleleles. In a single SMRT Sequencing run, scientists discovered 9 new alleles while generating fully-phased class II HLA genotypes for 48 samples3.
Explore this research further.
To learn more about how SMRT Sequencing powers imputation-free HLA typing, contact us.
- Hosomichi, K., et al., (2015) The impact of next-generation sequencing technologies on HLA research. Journal of Human Genetics. 60 (11), 665–673.
- Nelson, W. C., et al., (2015) An integrated genotyping approach for HLA and other complex genetic systems. Human Immunology. 76 (12), (928-938).
- Lang, K. et al. (September, 2015) Phased full-length SMRT Sequencing of HLA DPB1. Poster presented at 41st Annual Meeting of the American Society for Histocompatibility and Immunogenetics. Savannah, GA..
- Ashley, Euan A et al. (2016) Towards precision medicine. Nature Reviews. Genetics
- Gowda, Malali et al. (2016) Comparative analyses of low, medium and high-resolution HLA typing technologies for human populations Journal of Clinical & Cellular Immunology
- Shukla, Sachet A et al. (2015) Comprehensive analysis of cancer-associated somatic mutations in class I HLA genes. Nature Biotechnology
- Hosomichi, Kazuyoshi et al. (2015) The impact of next-generation sequencing technologies on HLA research. Journal of Human Genetics
- Mayor, Neema P et al. (2015) HLA typing for the next generation. PLoS One
- Kelleher, J et al. (2016) Identification of the novel B*27:144 allele in an Irish Individual. HLA
- Cereb, Nezih et al. (2015) Advances in DNA sequencing technologies for high resolution HLA typing. Human Immunology
- Hayward, D R et al. (2015) The novel HLA-B*44 allele, HLA-B*44:220, identified by Single Molecule Real-Time DNA sequencing in a British Caucasoid male. Tissue Antigens
- Hayhurst, J D et al. (2015) Two novel HLA alleles, HLA-A*30:02:01:03 and HLA-C*08:113, identified in a South African bone marrow donor. Tissue Antigens
- Chang, Chia-Jung et al. (2014) A fault-tolerant method for HLA typing with PacBio data. BMC Bioinformatics
- Poster: Pollard, M. O. et al. (2016) The MHC Diversity in Africa Project (MDAP) pilot – 125 African high resolution HLA types from 5 populations
- Poster: Cereb, N et al. (2016) Whole gene sequencing of KIR-3DL1 with SMRT Sequencing and the distribution of allelic variants in different ethnic groups
- Poster: Jenny Gu, Kevin Eng, Anand Sethuraman, Steve Kujawa et al. (2016) Multiplex target enrichment using barcoded multi-kilobase fragments and probe-based capture technologies
- Poster: Baybayan, Primo et al. (2016) Application specific barcoding strategies for SMRT Sequencing
- Poster: Ranade, Swati et al. (2016) Immune regions are no longer incomprehensible with SMRT Sequencing
- Poster: Pollard, Martin O. et al. (2015) HLA variant identification techniques
- Poster: Ranade, S. et al. (2015) Access full spectrum of polymorphisms in HLA class I & II genes, without imputation for disease association and evolutionary research.
- Poster: Lang, Kathrin et al. (2015) Full-length sequencing of HLA class I genes of more than 1000 samples provides deep insights into sequence variability
- Poster: Lang, Kathrin et al. (2015) Phased full-length SMRT Sequencing of HLA DPB1
- Poster: Ranade, Swati et al. (2015) Multiplexing human HLA class I & II genotyping with DNA barcode adapters for high throughput research.
- Poster: Mayor, Neema P et al. (2014) Genomic DNA sequences of HLA class I alleles generated using multiplexed barcodes and SMRT DNA Sequencing technology.
- Poster: Schöne, B. et al. (2016) Analysis of 37,000 Caucasian samples reveals tight linkage between SNP RS9277534 and high resolution typing of HLA-DPB1
- Poster: Cahill, Nicola et al. (2014) HLA sequencing using SMRT Technology – High resolution and high throughput HLA genotyping in a clinical setting
- Poster: Lleras, Roberto A et al. (2014) A novel analytical pipeline for de novo haplotype phasing and amplicon analysis using SMRT Sequencing technology.
- Cereb, Nezih (2017) AGBT PacBio Workshop: High-throughput HLA class I whole gene and HLA class II long range typing on PacBio RSII and Sequel Platforms
- (2017) Tutorial: Long Amplicon Analysis application
- Pollard, Martin (2016) ASHG Virtual Poster: The MHC Diversity in Africa Project (MDAP) pilot – 125 African high resolution HLA types from 5 populations
- Ashley, Euan (2016) AGBT Roche and PacBio Workshop: Towards precision medicine
- Ranade, Swati (2016) AGBT Virtual Poster: Immune regions are no longer incomprehensible with SMRT Sequencing
- Marsh, Steven (2015) Mendelspod: Long-read sequencing dramatically improves blood matching – Steven Marsh
- Mayor, Neema (2015) Seminar: Benefits of SMRT Sequencing for HLA typing at Anthony Nolan
- Hon, Lawrence and Raterman, Denise (2015) Xtalks Webinar: Long genomic DNA fragment capture and SMRT Sequencing enables accurate phasing of cancer and HLA loci
- Gyllensten, Ulf (2015) AGBT Virtual Poster: Clinical sequencing using Pacific Biosciences RS II for HLA typing and monitoring of drug resistance in chronic myeloid leukemia (CML)
- Marsh, Steven (2014) ASHI PacBio Workshop: The challenge of HLA diversity in 2014
- Tilgner, Hagen (2014) ASHG PacBio Workshop: Personalized and allele specific long-read transcriptomes
- Nagasaki, Masao (2015) ASHG Virtual Poster: Sequencing and typing of HLA class I genes by using a single molecule, real-time sequencing technology
- Cereb, Nezih (2014) Customer Experience: Histogenetics CEO, shares the reasons for adopting the PacBio sequencing platform
- Gyllensten, Ulf (2014) Mendelspod: The progress of clinical genomics in Sweden
- Korlach, Jonas (2014) Labroots Webinar: More comprehensive views of human genetic variation
- SMRT Sequencing Brochure: Revolutionize genomics with SMRT Sequencing (2017)
- Application Note: Multiplex target enrichment using barcoded multi-kilobase fragments and probe-based capture technologies (2016)
- HLA Sequencing Application Brochure: Fully phased, allele-specific HLA sequencing – the perfect pair (2015)
- Immunology Brochure: Invaluable insights into immunology (2015)
- Targeted Sequencing Application Brochure: Capture your regions of interest in high resolution (2015)
- Human Biomedical Research Brochure: The most comprehensive view of the human genome (2015)
- Product Note: Barcoded adapters and barcoded universal primers. (2015)
- Featured Interview: Revolutionize Translational Research – Uppsala’s Ulf Gyllensten on how long reads give access to new areas of the genome. (2015)
- Featured Interview: “We’re Going to Find the Keys” – Dan Geraghty discusses an approach to understanding causal genetic variation. (2014)
- Application Note: High-fidelity sequencing of complex genomic regions and full-length transcripts. (2014)