Long-read sequencing
The long-range PCR target enrichments of >20 ng were prepared for sequencing with the ONT ligation sequencing kit (SQK-LSK109) following the manufacturer’s protocol, with adjustments for sample type and yield. Individual sample libraries with <20 ng were concentrated at given bead clean-up steps. For sample pooling steps, samples were pooled in groups based on fragment size, to allow for more accurate normalisation prior to final pooling and flow cell loading. Prepared samples were sequenced on the MinION using the FLO-MIN106 version 9.4.1 flow cell platform. Flow cells were run until complete pore exhaustion, on average this was 72 hours, with minimal refuel of flow cells performed whenever active pore percentages dropped below 70%, achieving up to 30 billion basecall yields per flow cell and an average coverage depth per sample of >30,000x.