Abstract
Incontinentia pigmenti (IP) is a rare hereditary disorder affecting 1.2 in 100,000 live births, predominantly females. Genetic analysis of IP is complicated by a homologous pseudogene, making conventional short-read sequencing challenging. While long-range PCR is typically used to overcome this, skewed X-inactivation detection can also aid in assigning variants to IKBKG. We employed a comprehensive approach, incorporating whole-exome sequencing (WES), long-range PCR, RT-PCR, X-inactivation analysis, and nanopore sequencing, to identify and accurately phase a small heterozygous deletion, NM_001099857.5: c.363_367del, p.(Leu122Glyfs(∗)14), in the IKBKG gene in an IP-affected family. The deletion was initially detected via WES, with skewed X-inactivation observed in both the proband and her mother. Long-range PCR specific to IKBKG confirmed the variant's location in the IKBKG gene, not in the pseudogene. On the RNA level, the variant was undetectable, suggesting nonsense-mediated decay of the transcript. Nanopore sequencing precisely mapped the variant to IKBKG and analyzed the methylation status of both alleles, confirming the skewed X-inactivation, with the variant-carrying allele predominantly inactivated. This demonstrates the nanopore sequencing's value in genetic diagnosis, enabling precise variant localization and analysis of X chromosome activation status in females with skewed X-inactivation, aiding in accurate diagnosis and understanding of IP.