The Identification of Pompe Disease Mutations in Archival Tissues and Development of a Rapid Molecular-based Test
Aliya Alansari1, Samira Al-Rawahi, Taher Ba-Omar
1Department of Biology, College of Science, Sultan Qaboos University, Muscat, Oman.
Objectives:
Pompe disease (glycogen storage disease type II) is a rare autosomal recessive lysosomal storage disease that is caused by acid alpha-glucosidase deficiency. Early enzyme replacement therapy can benefit infants with the disease but the diagnosis is complicated by the rarity of the disease and the heterogeneity of the clinical manifestations. In this study, DNA extracted from archival postmortem formalin-fixed paraffin-embedded tissues was used to identify Pompe disease mutations in Oman and develop a rapid molecular-based test.
Methods:
Intronic primers were designed to amplify short fragments (193-454 base pairs [bp]) from coding exons (2-20) and screen for mutations using direct sequencing (DS).
Results:
Two mutations known to cause severe disease were identified in two samples. One was a coding mutation, c.2560C>T (p.Arg854X), and the second was found at a splice acceptor site, c.1327-2A>G. Polymerase chain reaction- and restriction fragment length polymorphism-based tests were designed for the rapid genotyping of the identified mutations.
Conclusion:
These tests can facilitate prenatal diagnosis and help in identifying carriers in families with the identified mutations.
Insights
Researchers identified two Pompe disease mutations in Oman using DNA from archival tissues. Rapid molecular tests were developed for genetic screening and carrier identification in affected families.
Area of Science:
- Genetics
- Molecular Biology
- Rare Diseases
Background:
- Pompe disease (glycogen storage disease type II) is a rare, autosomal recessive lysosomal storage disorder.
- Acid alpha-glucosidase deficiency underlies Pompe disease, impacting infants with early enzyme replacement therapy benefits.
- Diagnosis is challenging due to rarity and diverse clinical presentations.
Purpose of the Study:
- To identify Pompe disease mutations in Oman using DNA from archival postmortem tissues.
- To develop a rapid molecular-based diagnostic test for Pompe disease.
Main Methods:
- DNA extraction from formalin-fixed paraffin-embedded tissues.
- Direct sequencing of amplified coding exons (193-454 bp) using intronic primers.
- Development of PCR- and RFLP-based genotyping tests.
Main Results:
- Two known severe Pompe disease mutations were identified: c.2560C>T (p.Arg854X) and c.1327-2A>G.
- The identified mutations were located in a coding exon and a splice acceptor site.
- Rapid molecular tests were successfully designed for genotyping.
Conclusions:
- Developed tests facilitate rapid genotyping of identified Pompe disease mutations.
- These molecular tests can aid in prenatal diagnosis and carrier identification.
- The study provides a molecular tool for managing Pompe disease in Oman.
