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Apolipoproteins play a central role in cholesterol transport by their association to lipoproteins and their function as ligands for receptors, cofactors, or structural proteins. Mutations and polymorphisms in a variety of apolipoproteins lead to lipoprotein metabolism disorders and/or susceptibility to cardiovascular disease. In familial defective apolipoprotein B-100, the clearance of LDL particles from the circulation is impaired because of reduced affinity of the apolipoprotein (apo) B component of LDL for the LDL receptor as a result of a G-to-A mutation at nucleotide 10708 in exon 26 of the apoB gene, which causes substitution of Arg3500 for Gln (1)(2). The frequency of the mutation is 1 in 700 in the general population (3). Heterozygous individuals have increased serum concentrations of cholesterol. Apolipoprotein E (apoE) likewise is involved in the clearance of HDL, contributing to reverse cholesterol transport (4). Genetic variation at the APOE locus in humans is an important determinant of plasma lipid concentrations and relative risk of atherosclerosis (5). Among several rare variants, three major alleles have been identified in the population: E2 , E3 , and E4 . The most common E3 isoform is distinguished by cysteine at position 112 and arginine at position 158 in the receptor-binding region of apoE. The E4 isoform (Arg112 and Arg158) is associated with increased cholesterol, thus enhancing the risk of heart disease (5). In addition, E4 / E4 individuals have a very high risk for developing Alzheimer disease(6). Most patients with type III hyperlipidemia are homozygous for the E2 allele (Cys112 and Cys158).To date, genotyping for apoE is mainly performed by PCR, followed by digestion with restriction enzymes and restriction fragment length polymorphism (RFLP) analysis, and separation of the resulting DNA fragments on agarose or acrylamide gels (7). For apoB3500, an allele-specific …

High-Speed Apolipoprotein E Genotyping and Apolipoprotein B3500 Mutation Detection Using Real-Time Fluorescence PCR and Melting Curves