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Efficient Sequencing, Assembly, and Annotation of Human KIR Haplotypes

View ORCID ProfileDavid Roe, Jonathan Williams, Keyton Ivery, Jenny Brouckaert, Nick Downey, Chad Locklear, Rui Kuang, View ORCID ProfileMartin Maiers
doi: https://doi.org/10.1101/2020.07.12.199570
David Roe
1Bioinformatics and Computational Biology, University of Minnesota, Rochester, MN, USA
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  • For correspondence: roex0050@umn.edu
Jonathan Williams
2DNA Identification Testing Division, Laboratory Corporation of America Holdings, Burlington, NC, USA
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Keyton Ivery
2DNA Identification Testing Division, Laboratory Corporation of America Holdings, Burlington, NC, USA
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Jenny Brouckaert
2DNA Identification Testing Division, Laboratory Corporation of America Holdings, Burlington, NC, USA
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Nick Downey
3Integrated DNA Technologies, Inc. 1710 Commercial Park, Coralville, Iowa USA
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Chad Locklear
3Integrated DNA Technologies, Inc. 1710 Commercial Park, Coralville, Iowa USA
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Rui Kuang
1Bioinformatics and Computational Biology, University of Minnesota, Rochester, MN, USA
4Department of Computer Science and Engineering, Minneapolis, MN, USA
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Martin Maiers
5Center for International Blood and Marrow Transplant Research, Minneapolis, MN, USA
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Abstract

The homology, recombination, variation, and repetitive elements in the natural killer-cell immunoglobulin-like receptor (KIR) region has made full haplotype DNA interpretation impossible without physical separation of chromosomes. Here, we present a new approach using long-read sequencing to efficiently capture, sequence, and assemble diploid human KIR haplotypes. Sequences for capture probe design were derived from public full-length gene and haplotype sequences. IDT xGen® Lockdown probes were used to capture 2-8 kb of sheared DNA fragments followed by sequencing on a PacBio Sequel. The sequences were error corrected, binned, and then assembled using the Canu assembler. The assembly was evaluated on 16 individuals (8 African American and 8 Europeans) from whom ground truth was known via long-range sequencing on fosmid-isolated chromosomes. Using only 18 capture probes, the results show that the assemblies cover 97% of the GenBank reference, are 99.97% concordant, and it takes only 1.8 contigs to cover 75% of the reference. We also report the first assembly of diploid KIR haplotypes from long-read WGS, including the first sequencing of cB05∼tB01, which pairs a KIR2DS2/KIR2DS3 fusion with the tB01 region. Our targeted hybridization probe capture and sequencing approach is the first of its kind to fully sequence and phase all diploid human KIR haplotypes, and it is efficient enough for population-scale studies and clinical use.

Competing Interest Statement

JW, KI, and JB are or were employees of Laboratory Corporation of America Holdings. ND and CL are or were employees of Integrated DNA Technologies, Inc.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted July 13, 2020.
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Efficient Sequencing, Assembly, and Annotation of Human KIR Haplotypes
David Roe, Jonathan Williams, Keyton Ivery, Jenny Brouckaert, Nick Downey, Chad Locklear, Rui Kuang, Martin Maiers
bioRxiv 2020.07.12.199570; doi: https://doi.org/10.1101/2020.07.12.199570
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Efficient Sequencing, Assembly, and Annotation of Human KIR Haplotypes
David Roe, Jonathan Williams, Keyton Ivery, Jenny Brouckaert, Nick Downey, Chad Locklear, Rui Kuang, Martin Maiers
bioRxiv 2020.07.12.199570; doi: https://doi.org/10.1101/2020.07.12.199570

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