Simple and effective determination of apolipoprotein E genotypes by positive/negative polymerase chain reaction products

Davide Seripa, Emanuela Signori, Carolina Gravina, Maria Giovanna Matera, Monica Rinaldi, Vito M. Fazio

Research output: Contribution to journalArticle

26 Citations (Scopus)

Abstract

Several protein and DNA-based methods have been previously described for the identification of apolipoprotein E isoforms or genotypes. However, all of them generate frequently false-positive results. The purpose of this study was to set up a new, simple, and effective method for the analysis of the apoE polymorphism. A total of 1253 subjects previously examined for the apolipoprotein E polymorphism by restriction fragment length polymorphism were reanalyzed by our new method based on Taq DNA polymerase's inability to correctly initiate the replication in the presence of a mismatch at the 3′ end of the primer. We conceived a combination of 4 specific primers in 3 different pairs sharing the same stringent polymerase chain reaction conditions to directly detect the presence/absence of polymerase chain reaction products, and thus reveal the 6 apolipoprotein E genotypes. We confirm our previous results in 1171 subjects, whereas in 82 subjects out of 1253 (about 6%), the results have been reinterpreted. The final analysis revealed a total of 12 homozygotic subjects for the e2 allele (1.0%), 874 homozygotes for the e3 allele (69.8 %), and 8 homozygotes for the e4 allele (0.6 %). The frequence of heterozygotes was 8.7% for the e2/e3 genotype (n=109), 1.4% for the e2/e4 genotype (n=17), and 0.6% for the e3/e4 genotype (n=8). Relative allele frequencies were e2=0.060, e3=0.834, and e4=0.106. We describe a new, simple, unequivocal, and nonexpensive method for the identification of the 6 apoE genotypes.

Original languageEnglish
Pages (from-to)180-185
Number of pages6
JournalDiagnostic Molecular Pathology
Volume15
Issue number3
DOIs
Publication statusPublished - Sep 2006

Fingerprint

Apolipoproteins E
Genotype
Polymerase Chain Reaction
Alleles
Homozygote
Taq Polymerase
Heterozygote
Gene Frequency
Restriction Fragment Length Polymorphisms
Protein Isoforms
DNA
Proteins

Keywords

  • Allele-specific PCR
  • Apolipoprotein E
  • Genotyping
  • Polymorphism
  • Primer mismatch

ASJC Scopus subject areas

  • Pathology and Forensic Medicine

Cite this

Simple and effective determination of apolipoprotein E genotypes by positive/negative polymerase chain reaction products. / Seripa, Davide; Signori, Emanuela; Gravina, Carolina; Matera, Maria Giovanna; Rinaldi, Monica; Fazio, Vito M.

In: Diagnostic Molecular Pathology, Vol. 15, No. 3, 09.2006, p. 180-185.

Research output: Contribution to journalArticle

Seripa, Davide ; Signori, Emanuela ; Gravina, Carolina ; Matera, Maria Giovanna ; Rinaldi, Monica ; Fazio, Vito M. / Simple and effective determination of apolipoprotein E genotypes by positive/negative polymerase chain reaction products. In: Diagnostic Molecular Pathology. 2006 ; Vol. 15, No. 3. pp. 180-185.
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