Nanopore sequencing of the pharmacogene CYP2D6 allows simultaneous haplotyping and detection of duplications

CYP2D6 is one of the most important and widely studied genes in pharmacogenetics but it is highly polymorphic, can be affected by different forms of structural variation, and has a nearby pseudogene with highly similar sequence; therefore, its accurate genotyping and structural resolution are difficult to achieve with short sequencing reads. Liau and colleagues performed long-read amplicon sequencing of CYP2D6 on the GridION platform, which enabled its high-throughput genotyping, accurate haplotyping and variant detection.

Aim
Long-read sequencing offers the promise of overcoming some of the challenges in accurate genotyping of complex genes, along with the advantage of straightforward variant phasing. We have established methods for sequencing and haplotyping of the whole CYP2D6 gene using nanopore sequencing. 

Materials and methods
32 samples covering various haplotypes including gene duplication were sequenced on the GridION platform. 

Results
Haplotypes of 52 alleles matched accurately to known star (*) allele subvariants, with the remaining 12 being assigned as new alleles, or new subvariants of known alleles. Duplicated alleles could be detected by analyzing the allelic balance. 

Conclusion
Nanopore sequencing of CYP2D6 offers a high throughput method for accurate haplotyping, detection of new variants and determination of duplicated alleles.