Background Autosomal dominant polycystic kidney disease (ADPKD) is mainly caused by heterogeneous variants in the PKD1 and PKD2 genes. Genetic analysis of PKD1 has been challenging due to homology with 6 PKD1 pseudogenes and high GC content. Methods A single-tube multiplex long-range-PCR and long-read sequencing-based assay termed “comprehensive analysis of ADPKD” (CAPKD) was developed and evaluated in 170 unrelated patients by comparing to control methods including next-generation sequencing (NGS) and multiplex ligation-dependent probe amplification. Results CAPKD achieved highly specific analysis of PKD1 with a residual noise ratio of 0.05% for the 6 pseudogenes combined. CAPKD identified PKD1 and PKD2 variants (ranging from variants of uncertain significance to pathogenic) in 160 out of the 170 patients, including 151 single-nucleotide variants (SNVs) and insertion-deletion variants (indels), 6 large deletions, and one large duplication. Compared to NGS, CAPKD additionally identified 2 PKD1 variants (c.78_96dup and c.10729_10732dup). Overall, CAPKD increased the rate of variant detection from 92.9% (158/170) to 94.1% (160/170), and the rate of diagnosis with pathogenic or likely pathogenic variants from 82.4% (140/170) to 83.5% (142/170). CAPKD also directly determined the cis-/trans-configurations in 11 samples with 2 or 3 SNVs/indels, and the breakpoints of 6 large deletions and one large duplication, including 2 breakpoints in the intron 21 AG-repeat of PKD1, which could only be correctly characterized by aligning to T2T-CHM13. Conclusions CAPKD represents a comprehensive and specific assay toward full characterization of PKD1 and PKD2 variants, and improves the genetic diagnosis for ADPKD.

中文翻译：

---

常染色体显性多囊肾病长读长测序综合分析 PKD1 和 PKD2


背景 常染色体显性多囊肾病（ADPKD）主要由 PKD1 和 PKD2 基因的异质变异引起。由于 PKD1 与 6 个假基因的同源性和高 GC 含量，PKD1 的遗传分析一直具有挑战性。方法 通过与包括下一代测序 (NGS) 在内的对照方法进行比较，开发了一种基于单管多重长程 PCR 和长读长测序的检测方法，称为“ADPKD 综合分析”(CAPKD)，并在 170 名无关患者中进行了评估。和多重连接依赖性探针扩增。结果 CAPKD 实现了对 PKD1 的高度特异性分析，6 个假基因组合的残留噪声比为 0.05%。 CAPKD 在 170 名患者中的 160 名患者中鉴定出 PKD1 和 PKD2 变异（范围从不确定意义到致病性的变异），包括 151 个单核苷酸变异 (SNV) 和插入删除变异 (indels)、6 个大缺失和 1 个大重复。与 NGS 相比，CAPKD 还鉴定了 2 个 PKD1 变体（c.78_96dup 和 c.10729_10732dup）。总体而言，CAPKD 将变异检测率从 92.9% (158/170) 提高到 94.1% (160/170)，致病性或可能致病性变异的诊断率从 82.4% (140/170) 提高到 83.5% (142) /170）。 CAPKD还直接测定了11个具有2或3个SNV/indels的样本中的顺式/反式构型，以及6个大缺失和1个大重复的断点，其中2个断点位于PKD1的内含子21 AG重复中，仅能确定通过比对 T2T-CHM13 来正确表征。结论 CAPKD 代表了一种全面、特异性的检测方法，可全面表征 PKD1 和 PKD2 变异，并改善 ADPKD 的基因诊断。