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.
Park
Jason Y
JY
Department of Pathology and Laboratory Medicine, University of Pennsylvania School of Medicine and the Hospital of the University of Pennsylvania, Philadelphia, 19104, USA.
Narayan
Srinivas B
SB
Bennett
Michael J
MJ
D019943:Amino Acid Substitution; D001483:Base Sequence; D001681:Biological Assay / Q000379:methods*; D002334:Carnitine O-Palmitoyltransferase / Q000097:blood / Q000172:deficiency / Q000235:genetics*; D003596:Cytosine / Q000737:chemistry*; D005260:Female; D005838:Genotype; D006579:Heterozygote; D006801:Humans; D007223:Infant; D007930:Leucine / Q000235:genetics; D050356:Lipid Metabolism / Q000235:genetics / Q000502:physiology; D008297:Male; D017354:Point Mutation; D011392:Proline / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013941:Thymine / Q000737:chemistry*
D003596:Cytosine; D011392:Proline; D002334:Carnitine O-Palmitoyltransferase; D007930:Leucine; D013941:Thymine