A rapid and accurate methylation‐sensitive high‐resolution melting analysis assay for the diagnosis of Prader Willi and Angelman patients

Abstract

Abstract Background 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. Methods 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. Results 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. Conclusion The MS‐HRM showed good performance with a unique pair of primers, dispensing electrophoresis gel analysis, offering a quick and reproducible diagnostic.

Keywords

• Angelman syndrome
• high‐resolution melting
• MS‐HRM
• MS‐PCR
• Prader Willi syndrome