Toye Emuejevoke T, van Marle Guido, Hutchins Wendy, Abgabiaje Olayinka, Okpuzor Joy
University of Lagos, Department of Cell Biology and Genetics.
University of Calgary Cumming School of Medicine, Department of Microbiology Immunology and Infectious Diseases.
Afr Health Sci. 2018 Dec;18(4):995-1002. doi: 10.4314/ahs.v18i4.20.
Sickle cell anaemia (SCA) is a hereditary blood disorder caused by a single mutation in the haemoglobin gene. The disease burden of SCA is highest in Nigeria. The allele specific polymerase chain reaction (ASPCR) method is applicable for the direct detection of known single nucleotide polymorphisms (SNPs).
To investigate the use of the single tube ASPCR as an accurate and affordable method for SCA screening in Nigeria.
DNA was extracted from study subjects with normal haemoglobin, HbAA (20), sickle cell anaemia, HbSS (20) and carriers, HbAS (1). Haemoglobin was genotyped by ASPCR using two primer sets that amplifies the wildtype and mutant haemoglobins in each sample. Amplicon sizes were analyzed by gel electrophoresis.
Amplicons were visible after electrophoresis at regions 517 base pair (bp) for HbA and 267 bp for HbS. ASPCR correctly and unambiguously detected the presence or absence of haemoglobins A and S from all samples collected, demonstrating its accuracy and precision for the screening of SCA.
This study validates ASPCR as an effective, low cost approach for the clinical screening of SCA in Nigeria. ASPCR is also applicable for other genetic diseases, paternity testing, and forensics where more expensive fluorescence-based approaches are not obtainable.
镰状细胞贫血（SCA）是一种由血红蛋白基因突变引起的遗传性血液疾病。尼日利亚的SCA疾病负担最高。等位基因特异性聚合酶链反应（ASPCR）方法适用于直接检测已知的单核苷酸多态性（SNP）。
研究单管ASPCR作为尼日利亚SCA筛查的一种准确且经济实惠的方法的应用情况。
从血红蛋白正常的研究对象（HbAA，20例）、镰状细胞贫血患者（HbSS，20例）和携带者（HbAS，1例）中提取DNA。使用两组引物通过ASPCR对血红蛋白进行基因分型，这两组引物可扩增每个样本中的野生型和突变型血红蛋白。通过凝胶电泳分析扩增子大小。
电泳后，在517碱基对（bp）处可见HbA的扩增子，在267 bp处可见HbS的扩增子。ASPCR正确且明确地检测出了所有采集样本中血红蛋白A和S的存在与否，证明了其在SCA筛查中的准确性和精确性。
本研究验证了ASPCR作为尼日利亚SCA临床筛查的一种有效、低成本方法的有效性。ASPCR也适用于其他遗传疾病、亲子鉴定以及无法获得更昂贵的基于荧光方法的法医鉴定。
