Cas9 targeted nanopore sequencing with enhanced variant calling improves CYP2D6-CYP2D7 hybrid allele genotyping

The study aimed to develop a more accurate method for genotyping the highly polymorphic CYP2D6 gene, which is challenging due to its similarity to neighbouring pseudogenes like CYP2D7 and the frequent occurrence of hybrid alleles. These genetic variations are critical for drug metabolism. The researchers focused on optimising nanopore Cas9-targeted sequencing (nCATS) combined with adaptive sequencing to generate complete allele-specific consensus sequences, improving the detection of both large structural variants and small variants that are typically missed by conventional genotyping assays.

The Oxford Nanopore MinION sequencer was used in this study, with the sequencing technique enhanced by Cas9-targeted sequencing to enrich specific genomic regions of interest. A newly developed analysis pipeline, CoLoRGen, was used to detect variants and assign accurate star alleles for the CYP2D6 and CYP2D7 genes.

The major findings revealed that this optimized method could accurately identify CYP2D6-CYP2D7 hybrid alleles, including novel structural variants that were not detectable by current genotyping methods. The researchers demonstrated that their approach could improve genotyping accuracy by phasing de novo mutations and providing more accurate gene function predictions. This enhanced genotyping method could be instrumental in improving pharmacogenetic profiling for clinical applications, enabling better personalised medicine.