【摘要】 目的 探讨通过长读长测序技术构建SNP单体型，为成人常显多囊肾基因（PKD1）致病变异且家系不全患者行胚胎植入前遗传学检测策略提供参考。方法 选取2022年1月至2022年9月于柳州市妇幼保健院生殖健康助孕中心就诊的两名患者，分别患有PKD1 c.5976＿5978del杂合变异复合PKD1c.11333C>A杂合变异和PKD1 c.11969 T>G杂合变异导致的常染色体显性遗传多囊肾且家系不全，采集患者夫妻双方外周血提取高质量DNA，通过长读长测序技术构建SNP单体型。活检胚胎滋养外胚层细胞进行全基因组扩增，再利用ASA芯片检测胚胎SNP位点信息，结合长读长测序构建的男女方SNP位点建立连锁分析，分析胚胎变异位点携带情况；同时以Sanger测序检出的受累胚胎作为先证者，利用Karyomapping芯片检测分析胚胎SNP位点单体型；比较两种连锁分析结果的一致性。结果 病例1检测5个胚胎，病例2送检4个胚胎。病例1的1号，3号和4号胚胎携带男方PKD1 c.5976＿5978del致病变异，病例2的1号及4号胚胎均携带有男方PKD1 c.11969T>G致病变异；经Sanger测序均选择1号胚胎作为参照，利用Karyomapping芯片进行连锁分析，病例1的1号，3号和4号，病例2的1号及4号胚胎均携带男方的致病变异，结果与长读长测序家系连锁分析一致。结论 对于家系不全PKD1患者，可通过长度长测序进行男女双方的致病位点确定及SNP单体型构建，通过SNP单体型建立连锁分析，从而准确有效的进行胚胎植入前遗传学检测。更多还原

【Abstract】 Objective To explore the construction of single nucleotide polymorphism(SNP) haplotype based on the technology of long-read sequencing, to provide reference for preimplantation genetic testing strategies for patients with adult polycystic kidney disease(PKD1) pathogenic mutations and incomplete pedigrees. Methods Two patients visited the Reproductive Health Assisted Pregnancy Center of Liuzhou Maternal and Child Health Hospital from January 2021 to September 2022. Both patients had autosomal dominant polycystic kidney disease caused by gene variations of PKD1 c.5976＿5978del, PKD1 c.11333C>A and PKD1c. 11969 T>G heterozygous mutations(with incomplete pedigree). Peripheral blood samples were collected from the husband and wife of these patients to extract high-quality DNA. SNP haplotypes were constructed using long-read sequencing technology. Biopsies of trophoblastic ectodermal cells were performed for whole-ge-nome amplification, followed by the detection of embryo SNP site information using the Illumina Asian Screening Array(ASA). Linkage analysis was then established by combining the SNP sites of males and females constructed by long-read sequencing to analyze embryo mutations. Simultaneously, affected embryos detected by Sanger sequencing were used as probands for Karyomapping to detect and analyze embryo SNP haplotypes Finally, the results of the two linkage analyses were compared for consistency. Results Five embryos were tested for Case 1, and four embryos for Case 2. In case 1, embryos 1, 3, and 4 carried the PKD1 c.5976＿5978del pathogenic variant from the male side. In case 2, embryos 1 and 4 carried the PKD1 c.11969T>G pathogenic variant from the male side. After Sanger sequencing, Karyomapping chips were used for linkage analysis. Using the affected embryo 1 as the reference. It was found that embryos 1, 3, and 4 of case 1, and embryos 1 and 4 of case 2 all carried the pathogenic variant from the male side. This was consistent with the linkage analysis results from the long-read sequencing family. Conclusion For patients with PKD1 pathogenic mutations and incomplete pedigrees, long-read sequencing may help determine the pathogenic sites in both males and females. Additionally, it can aid in constructing SNP haplotypes. Subsequent linkage analysis can then be performed based on the constructed SNP haplotypes, leading to accurate and effective PGT.更多还原