Laboratório de Alta Complexidade, Instituto Nacional da Saúde da Mulher, da Criança e do Adolescente Fernandes Figueira, Fiocruz, Rio de Janeiro, Brazil.
Departamento de Genética, Instituto Nacional da Saúde da Mulher, da Criança e do Adolescente Fernandes Figueira, Fiocruz, Rio de Janeiro, Brazil.
Mol Genet Genomic Med. 2019 Jun;7(6):e637. doi: 10.1002/mgg3.637. Epub 2019 Apr 29.
Prader Willi (PWS) and Angelman (AS) syndromes are rare genetic disorders characterized by deletions, uniparental disomy, and imprinting defects at chromosome 15. The loss of function of specific genes caused by genetic alterations in paternal allele causes PWS while the absence in maternal allele results AS. The laboratory diagnosis of PWS and AS is complex and demands molecular biology and cytogenetics techniques to identify the genetic mechanism related to the development of the disease. The DNA methylation analysis in chromosome 15 at the SNURF-SNRPN locus through MS-PCR confirms the diagnosis and distinguishes between PWS and AS. Our study aimed to establish the MS-PCR technique associated with High-Resolution Melting (MS-HRM) in PWS and AS diagnostic with a single pair of primers.
We collected blood samples from 43 suspected patients to a cytogenetic and methylation analysis. The extracted DNA was treated with bisulfite to perform comparative methylation analysis.
MS-HRM and MS-PCR agreed in 100% of cases, identifying 19(44%) PWS, 3(7%) AS, and 21(49%) Normal. FISH analysis detected four cases of PWS caused by deletions in chromosome 15.
The MS-HRM showed good performance with a unique pair of primers, dispensing electrophoresis gel analysis, offering a quick and reproducible diagnostic.
普拉德-威利（PWS）和安格曼（AS）综合征是罕见的遗传疾病，其特征是染色体 15 上的缺失、单亲二体和印迹缺陷。由于父本等位基因的遗传改变导致特定基因功能丧失引起 PWS，而母本等位基因缺失则导致 AS。PWS 和 AS 的实验室诊断较为复杂，需要分子生物学和细胞遗传学技术来确定与疾病发展相关的遗传机制。通过 MS-PCR 在染色体 15 的 SNURF-SNRPN 基因座进行 DNA 甲基化分析可确认诊断，并区分 PWS 和 AS。我们的研究旨在建立一种单对引物的 MS-PCR 技术，用于 PWS 和 AS 的诊断。
我们从 43 名疑似患者中采集血液样本进行细胞遗传学和甲基化分析。提取的 DNA 经亚硫酸氢盐处理后进行比较甲基化分析。
MS-HRM 和 MS-PCR 在 100%的病例中一致，确定了 19 例（44%）PWS、3 例（7%）AS 和 21 例（49%）正常。FISH 分析检测到 4 例由染色体 15 缺失引起的 PWS。
MS-HRM 表现出良好的性能，使用独特的一对引物，无需电泳凝胶分析，提供快速且可重复的诊断。
