Apolipoprotein E genotyping method by real time PCR, a fast and cost-effective alternative to the TaqMan and FRET assays

Olga Calero; Rafael Hortigüela; María J Bullido; Miguel Calero

doi:10.1016/j.jneumeth.2009.06.033

Apolipoprotein E genotyping method by real time PCR, a fast and cost-effective alternative to the TaqMan and FRET assays

J Neurosci Methods. 2009 Oct 15;183(2):238-40. doi: 10.1016/j.jneumeth.2009.06.033. Epub 2009 Jul 5.

Authors

Olga Calero¹, Rafael Hortigüela, María J Bullido, Miguel Calero

Affiliation

¹ Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Spain.

PMID: 19583979
DOI: 10.1016/j.jneumeth.2009.06.033

Abstract

The apolipoprotein E gene (APOE) polymorphism genotyping has an allegedly important predictive value for coronary heart disorders and Alzheimer's disease. We developed a simple, fast, cost-effective and suited for high-throughput protocol for determining APOE genotypes by Real Time PCR monitored by SYBR Green. The method is based on differential amplification by allele-specific primers. These primers have variations in their 3'-end nucleotides such that are specific for one of the two variants in each polymorphic position. By this protocol, we obtained a 100% concordance with the APOE genotypes determined by sequencing analysis. The main advantages of this method are its relative simplicity and the reduced cost compared to other methodologies, such as the TaqMan and FRET assays.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Apolipoproteins E / genetics*
Fluorescence Resonance Energy Transfer / methods*
Genotype*
Humans
Polymerase Chain Reaction / methods*
Polymorphism, Single Nucleotide / genetics
Taq Polymerase*

Substances

Apolipoproteins E
Taq Polymerase