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A multiplex pharmacogenetics assay using the MinION nanopore sequencing device

Liau, Yusmiatia; Cree, Simone L.a; Maggo, Simrana; Miller, Allison L.a; Pearson, John F.a; Gladding, Patrick A.b; Kennedy, Martin A.a

doi: 10.1097/FPC.0000000000000385
Original Articles

Objectives The MinION nanopore sequencing device opens the opportunity to cost-effective and point-of-care DNA sequencing. As a proof of principle, we developed a multiplex assay targeting pharmacogenetic variants related to clopidogrel and warfarin, the two commonly used drugs that show response variability due to genetic polymorphisms.

Methods Six reference and 78 clinical DNA samples were amplified by PCR to generate 15 amplicons targeting 27 key variants. These products were then barcoded to enable sample multiplexing in one sequencing run. Four variant calling tools (marginCaller, VarScan 2, nanopolish, Clairvoyante) were used to compare genotyping accuracy.

Results In our cohort, 81 out of 84 samples were successfully sequenced and genotyped. Using nanopolish as the variant calling tool achieved accuracy >95% for all except two variants. A known single base deletion (CYP2C9*6) was successfully detected.

Conclusion While minor misgenotyping issues exist, this work demonstrates that drug-specific or broad pharmacogenetic screening assays using small PCR amplicons are possible on the MinION sequencing device.

aDepartment of Pathology and Biomedical Science, University of Otago, Christchurch

bTheranostics Laboratory, North Shore Hospital, Shakespeare Rd, Auckland, New Zealand

Received 4 June 2019 Accepted 4 August 2019

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Correspondence to Martin A. Kennedy, Department of Pathology and Biomedical Science, University of Otago, 2 Riccarton Avenue, Christchurch, New Zealand, Tel: +6433640590; fax: + 6433640009; e-mail:

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