Email updates

Keep up to date with the latest news and content from BMC Medical Genetics and BioMed Central.

Open Access Highly Accessed Research article

Allele-specific polymerase chain reaction for the detection of Alzheimer’s disease-related single nucleotide polymorphisms

Mohd Nazif Darawi1*, Chin Ai-Vyrn2, Kalavathy Ramasamy3, Philip Poi Jun Hua2, Tan Maw Pin2, Shahrul Bahyah Kamaruzzaman2 and Abu Bakar Abdul Majeed14

Author Affiliations

1 Brain Science Research Laboratory, Faculty of Pharmacy, Universiti Teknologi MARA, Puncak Alam, Selangor, 42300, Malaysia

2 Ageing and Age Associated Disorders Research Group, Department of Medicine, Faculty of Medicine, University Malaya, Kuala Lumpur, 50603, Malaysia

3 Collaborative Drug Discovery Research Group, Faculty of Pharmacy, Universiti Teknologi MARA, Puncak Alam, Selangor, 42300, Malaysia

4 Research Management Institute, Universiti Teknologi MARA, Shah Alam, Selangor, 40450, Malaysia

For all author emails, please log on.

BMC Medical Genetics 2013, 14:27  doi:10.1186/1471-2350-14-27

Published: 19 February 2013

Abstract

Background

The incidence of Alzheimer’s disease, particularly in developing countries, is expected to increase exponentially as the population ages. Continuing research in this area is essential in order to better understand this disease and develop strategies for treatment and prevention. Genome-wide association studies have identified several loci as genetic risk factors of AD aside from apolipoprotein E such as bridging integrator (BIN1), clusterin (CLU), ATP-binding cassette sub-family A member 7 (ABCA7), complement receptor 1 (CR1) and phosphatidylinositol binding clathrin assembly protein (PICALM). However genetic research in developing countries is often limited by lack of funding and expertise. This study therefore developed and validated a simple, cost effective polymerase chain reaction based technique to determine these single nucleotide polymorphisms.

Methods

An allele-specific PCR method was developed to detect single nucleotide polymorphisms of BIN1 rs744373, CLU rs11136000, ABCA7 rs3764650, CR1 rs3818361 and PICALM rs3851179 in human DNA samples. Allele-specific primers were designed by using appropriate software to permit the PCR amplification only if the nucleotide at the 3’-end of the primer complemented the base at the wild-type or variant-type DNA sample. The primers were then searched for uniqueness using the Basic Local Alignment Search Tool search engine.

Results

The assay was tested on a hundred samples and accurately detected the homozygous wild-type, homozygous variant-type and heterozygous of each SNP. Validation was by direct DNA sequencing.

Conclusion

This method will enable researchers to carry out genetic polymorphism studies for genetic risk factors associated with late-onset Alzheimer’s disease (BIN1, CLU, ABCA7, CR1 and PICALM) without the use of expensive instrumentation and reagents.

Keywords:
Alzheimer’s disease; Single nucleotide polymorphism; Apolipoprotein E; Bridging integrator; Clusterin; ATP-binding cassette sub-family A member 7; Complement receptor 1; Phosphatidylinositol binding clathrin assembly protein; Allele-specific polymerase chain reaction