The autosomal dominant form of polycystic kidney disease (ADPKD) is a hereditary kidney disease that results in late-onset renal cyst development, multisystem disorder, and possibly end-stage renal disease (ESRD) after the fifth decade of life[1]. With an estimated incidence of 1:1000 to 1:400 individuals, ADPKD is the most common single-gene inherited kidney disease primarily caused by pathogenic mutations in PKD1 and PKD2 genes, encoding polycystin-1 and polycystin-2, respectively[2–4]. Among these, approximately 85% of cases are caused by pathogenic mutations in PKD1 and about 15% by the PKD2 gene[5, 6]. As an autosomal dominant disease, ADPKD patients could pass on the pathogenic mutations to their offspring with a 50% chance, thus placing a heavy burden on families and the society suffering from the disease[7].
To prevent the transmission of pathogenic variants, preimplantation genetic testing for monogenic diseases (PGT-M) is increasingly used in clinical practice [8]. Theoretically, PGT-M is available for any monogenic disorder with identified disease-causing locus [9]. In a general PGT-M cycle, multiplex PCR for genetic markers followed by next-generation sequencing (NGS) or SNP array was applied to related family members to identify informative genetic markers for further haplotyping analysis[8, 9]. The high-risk haplotype linked with the familial pathogenic variant should be determined before the clinical cycle, especially in cases for PKD1 PGT-M where direct detection of mutations at the single-cell level is challenging because of homologous pseudogenes and high GC content. However, in some cases, it is hard to determine the high-risk haplotype for de novo pathogenic variant or no probands are available. For ADPKD, an estimated 10–15% of cases are de novo mutations or gonad mosaicism in parents[1]. Therefore, establishing a reliable linkage analysis for a prospective parent with a de novo mutation in PKD1 is essential in their PGT-M application.
Different from short-read-based NGS, third-generation sequencing (TGS) represented by PacBio and Oxford nanopore sequencing technologies (ONT) could generate long reads to cover more genomic regions, which is potential for direct haplotype phasing[10–13]. Moreover, TGS technologies can directly read the highly repetitive and high GC regions for more sequences[14].
In this study, we reported a successful application of ONT-based long-read sequencing for a female patient seeking PGT-M with de novo PKD1 mutation and successfully deduced the high-risk haplotype. Based on the direct haplotype results and targeted SNP for the couple and the six embryos in the clinical cycle, we identified the carrier status for these embryos. The selected embryo was transplanted and led to the birth of a healthy baby without the inherited mutation in PKD1. The result showed the potential of long-read sequencing for direct reconstruction of individual haplotypes, especially for de novo mutation carriers, and facilitate identification of embryo status in PGT-M application. TGS for direct haplotyping together with NGS was proven to be a practical and promising strategy for PGT-M involved with de novo mutation or incomplete pedigree.