title: Molecular assay of -alpha(3.7) and -alpha(4.2) deletions causing alpha-thalassemia by denaturing high-performance liquid chromatography.
abstract: OBJECTIVE
  alpha-Thalassemia, the most common single gene disorder in humans, is due to the absence of one (-alpha/alphaalpha) or both (--/alphaalpha) of the two functional alpha-globin genes (alpha1 and alpha2). The -alpha(3.7) and -alpha(4.2) single gene deletions are common in Southeast Asian populations. Southern blotting analysis and gap PCR assay are commonly used for the detection of such alpha-thalassemia genotypes. The two genes are located on chromosome 16, with high homology (>96%).
  
  
  METHODS
  Based on the sequence variation within the two Z boxes, a denaturing high-performance liquid chromatography (DHPLC)-based assay was developed for rapid genotyping of the -alpha(3.7) and -alpha(4.2) alleles. To demonstrate the utility of this approach, 40 DNA samples with known genotypes were analyzed, including -alpha(3.7)/alphaalpha (7 cases), -alpha(4.2)/alphaalpha (4 cases), -alpha(3.7)/--(SEA) (6 cases), -alpha(4.2)/--(SEA) (3 cases), and 20 unaffected subjects (alphaalpha/alphaalpha).
  
  
  RESULTS
  We successfully distinguished all of the alpha-thalassemia genotypes through their characteristic chromatograms of alpha1 and alpha2 genes. The accuracy of this technique for our sample was 100% sensitivity and specificity.
  
  
  CONCLUSIONS
  This novel and alternative DHPLC-based alpha-thalassemia genotype assay is easy, rapid, and highly accurate. This technique enables the diagnosis of silent alpha+ thalassemia and hemoglobin H disease for large scale population screening.
authors.affiliation: Institute of Biomedical Engineering, College of Medicine and College of Engineering, National Taiwan University, Taipei, Taiwan.
authors.forename: Chia-Cheng
authors.initials: CC
authors.lastname: Hung
authors.forename: Chien-Nan
authors.initials: CN
authors.lastname: Lee
authors.forename: Chih-Ping
authors.initials: CP
authors.lastname: Chen
authors.forename: Yuh-Jyh
authors.initials: YJ
authors.lastname: Jong
authors.forename: Wu-Shiun
authors.initials: WS
authors.lastname: Hsieh
authors.forename: Win-Li
authors.initials: WL
authors.lastname: Lin
authors.forename: Yi-Ning
authors.initials: YN
authors.lastname: Su
authors.forename: Su-Ming
authors.initials: SM
authors.lastname: Hsu
mesh_terms: D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D017353:Gene Deletion; D005838:Genotype; D006801:Humans; D008969:Molecular Sequence Data; D009691:Nucleic Acid Denaturation; D017085:alpha-Thalassemia / Q000235:genetics*