The advantages of haplotype analysis of the promoter region of the human apolipoprotein E gene.
Polymorphisms in the regulatory region of the human apolipoprotein E gene (gene, APOE; protein, apoE) have been implicated in Alzheimer's disease. Here we describe in detail the advantages of a simple method for haplotype analysis of this region (at -491 and -427 bases relative to the transcription start site of the gene). The promoter region of the APOE gene was amplified by polymerase chain reaction (PCR) and this fragment was then used as a template for PCR with "nested" primers to generate a 228-bp product incorporating both the -491 and the -427 loci. PCR products were then digested with DraI and AluI together and subjected to polyacrylamide gel electrophoresis. The distinct pattern of bands appearing on the gel was then used to ascribe [-491,-427] haplotypes to each subject, from which -491 and -427 genotypes were inferred. -491 and -427 genotypes were also confirmed by digestion with DraI alone or AluI alone. Haplotype analysis was successful in all 20 samples analyzed and was 100% consistent with genotyping. We suggest that this is a reliable, time-saving method that the will be useful in large-scale APOE promoter genotyping studies.
McLaughlin
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Academic Department of Anaesthesia and Intensive Care, Barts, Queen Mary, University of London, The Royal London Hospital, Whitechapel, London E1 1BB, United Kingdom.
Sharma
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McGinley
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Samra
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D000328:Adult; D001057:Apolipoproteins E / Q000235:genetics*; D005260:Female; D006239:Haplotypes / Q000235:genetics*; D006801:Humans; D008297:Male; D008875:Middle Aged; D016133:Polymerase Chain Reaction / Q000379:methods*; D011110:Polymorphism, Genetic; D011401:Promoter Regions, Genetic
D001057:Apolipoproteins E