Molecular assay for detection of the common carnitine palmitoyltransferase 1A 1436(C> T) mutation

Background: Carnitine palmitoyltransferase 1A (CPT1A) deficiency is a metabolic disorder that occurs at a key checkpoint of fatty acid metabolism. A new form of CPT1A deficiency caused by a mutation at nucleotide 1436 (C > T), resulting in an amino acid substitution of leucine for proline at position 479 (P479L), has been isolated in Canadian First Nations and Inuit populations. The present study offers a molecular method for assessing CPT1A 1436 (C > T) mutation status. Methods: CPT1A-deficient fibroblasts from four patient fibroblast cell lines and ten patient peripheral blood spots were all analyzed by polymerase chain reaction (PCR) coupled to restriction endonuclease ( RE) treatment. Genomic DNA was PCR-amplified and treated with an RE specific for normal DNA. CPT1A 1436 (C > T) mutations were identified by resistance to RE treatment. Results: The RE-PCR assay identified homozygosity for the 1436 (C > T) mutation in four fibroblast cell lines and nine blood spots with CPT1A enzyme deficiency. In addition, the assay identified one blood spot that corresponded to the heterozygous genotype. Conclusions: RE-PCR assay for the 1436 (C > T) mutation provides a rapid assay for the diagnosis of CPT1A deficiency resulting from this mutation. The assay will have utility in screening populations with a high prevalence of this genotype.