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

BMC Medical Genetics

Abu Bakar Abdul Majeed³  Shahrul Bahyah Kamaruzzaman¹  Tan Maw Pin¹  Philip Poi Jun Hua¹  Kalavathy Ramasamy²  Chin Ai-Vyrn¹  Mohd Nazif Darawi⁴ 

[1] Ageing and Age Associated Disorders Research Group, Department of Medicine, Faculty of Medicine, University Malaya, Kuala Lumpur, 50603, Malaysia;Collaborative Drug Discovery Research Group, Faculty of Pharmacy, Universiti Teknologi MARA, Puncak Alam, Selangor, 42300, Malaysia;Research Management Institute, Universiti Teknologi MARA, Shah Alam, Selangor, 40450, Malaysia;Brain Science Research Laboratory, Faculty of Pharmacy, Universiti Teknologi MARA, Puncak Alam, Selangor, 42300, Malaysia

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

Others  :  1177727
DOI  :  10.1186/1471-2350-14-27

 received in 2012-08-15, accepted in 2013-02-15,  发布年份 2013

【 摘 要 】

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.

【 授权许可】

   
2013 Darawi et al; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

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