Cas9 targeted nanopore sequencing with enhanced variant calling improves CYP2D6–CYP2D7 hybrid allele genotyping
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CYP2D6 is one of the most challenging pharmacogenes to genotype due to the high similarity with its neighboring pseudogenes and the frequent occurrence of CYP2D6-CYP2D7 hybrids. Unfortunately, most current genotyping methods are therefore unable to correctly determine the complete CYP2D6-CYP2D7 sequence. With this in mind, we developed a genotyping assay to generate complete allele-specific consensus sequences of complex regions, by optimising the PCR-free nanopore Cas9-targeted sequencing (nCATS) method combined with adaptive sequencing, and developing a new comprehensive long read genotyping (CoLoRGen) pipeline. In contrast to state-of-the-art variant callers, CoLoRGen first generates consensus sequences of both alleles and subsequently determines both large structural and small variants to ultimately assign the correct star-alleles.