High-resolution dissociation analysis for the molecular diagnosis of cystic fibrosis in Cuba.
DOI:
https://doi.org/10.56294/sctconf20251757Keywords:
CFTR, cystic fibrosis, HRM, sequencingAbstract
Cystic fibrosis is a common recessive disease in the Caucasian population. Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene can cause damage to the protein it encodes. Seven mutations are directly analyzed in Cuba, representing 55.5% of the pathogenic allelic variants in cystic fibrosis patients. The high allelic heterogeneity of this gene and its varied phenotypic expression make it vitally important to detect a greater number of genetic variants that cause the disease. With this data, patients can receive better genetic counseling and personalized treatment that would improve their quality of life. It is therefore necessary to search for new variants to complete the diagnosis. The objective of this work was to identify new genetic variants in the CFTR gene and confirm the clinical diagnosis of Cystic Fibrosis. Twenty-one patients clinically diagnosed as cystic fibrosis by the National Medical Genetics Network or identified as positive through the Neonatal Screening Program were analyzed. Eleven real-time polymerase chain reaction assays were performed on the isolated DNA, followed by high-resolution dissociation curve analysis. The amplification of these eleven fragments covered the five exons of the gene with the highest number of described mutations and the adjacent intronic regions. Subsequently, the samples that showed changes in the curve pattern were sequenced. Five mutations were detected: one pathogenic, two of uncertain significance, one benign, and one with conflicting clinical significance. Point genetic variants were identified in four of the five exons analyzed. The real-time polymerase chain reaction – high resolution melting curve technique proved useful for searching for point mutations along the CFTR gene sequence, which, in combination with Sanger sequencing, will allow for the identification of new variants in the Cuban population.
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Copyright (c) 2025 Teresa Collazo Mesa, Ixchel López Reyes, Mercy Estévez Miere, Alejandro Esperón Álvarez, Fidel Rodríguez Cala (Author)
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