fig5

Figure 5. Comparison of exome sequencing (ES), short-read and long-read genome sequencing (GS) in resolving complex regions (“dark regions”) of the human genome. An example of such regions is the PKD1 (polycystic kidney disease 1) gene, where the first 32 exons are located in a segmental duplicated region on chromosome 16p13, with six pseudogenes located 13 Mb proximal to the PKD1 locus. In addition to high GC content, the sequences of these six pseudogenes are highly homologous to PKD1 and share 97% sequence similarity, making amplification- and capture-based approaches challenging. The PKD1 region is visualised with Integrative Genomics Viewer (IGV) using different sequencing approaches. Despite improvements in the capture probe design, ES of exons 1 to 14 of PKD1 showed lower coverage, while GS achieved a more uniform coverage for the entire locus, including the duplicated region. Long-read GS enables unambiguous alignment of reads, complementing short-read GS, and enhances disease diagnosis. The orange double arrow indicates the “dark region”. The red dotted box and arrow indicate regions where short-read GS covers poorly.