title: A quantitative assay to detect α-thalassemia deletions and triplications using multiplex nested real-time quantitative polymerase chain reaction.
abstract: Increasing evidence indicates that copy number variants (CNVs) have great relevance to common human diseases. In α-thalassemia, clinical phenotypes are related to genotypes, specifically copy number changes in the human α-globin gene cluster. Assays are available for high-throughput screening of unknown CNVs genome-wide and also for targeted CNV genotyping at loci associated with genetic disorders. Here we describe a universal quantitative approach based on nested real-time quantitative polymerase chain reaction for accurate determination of copy numbers at multiple particular gene loci. We used the α-globin gene as a model system, obtaining the reproducibility and sensitivity to analyze different gene copies and testing 95 DNA samples with 16 different known genotypes. Our results showed that this approach has high sensitivity and low standard deviations for correctly genotyping DNA samples containing different copy numbers of the α1 and α2 globin genes. Our method is rapid, simple, and reliable, and it could be used to simultaneously screen for α-thalassemia deletions or triplications. Moreover, it has potential as a versatile technology for the rapid genotyping of known CNVs in a targeted region.
authors.affiliation: Department of Medical Genetics, School of Basic Medical Sciences, Southern Medical University, Guangzhou 510515, People's Republic of China.
authors.forename: Wan-Jun
authors.initials: WJ
authors.lastname: Zhou
authors.forename: Wei-Yu
authors.initials: WY
authors.lastname: Liu
authors.forename: Dun
authors.initials: D
authors.lastname: Liu
authors.forename: Jian-Hui
authors.initials: JH
authors.lastname: Jiang
authors.forename: Dian-Min
authors.initials: DM
authors.lastname: Zhou
authors.forename: Ze-Yan
authors.initials: ZY
authors.lastname: Zhong
authors.forename: Xiang-Min
authors.initials: XM
authors.lastname: Xu
mesh_terms: D001483:Base Sequence; D056915:DNA Copy Number Variations; D016172:DNA Fingerprinting / Q000379:methods*; D005784:Gene Amplification; D018628:Gene Dosage; D056426:Genetic Loci; D005838:Genotype; D006801:Humans; D008969:Molecular Sequence Data; D060885:Multiplex Polymerase Chain Reaction; D009154:Mutation; D020033:Protein Isoforms / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D017384:Sequence Deletion; D055542:alpha-Globins / Q000235:genetics*; D017085:alpha-Thalassemia / Q000235:genetics*
chemical_list: D020033:Protein Isoforms; D055542:alpha-Globins