title: Direct genotyping and prenatal diagnosis of beta-thalassemia in Chinese by polymerase chain reaction mediated restriction fragment length polymorphism method.
abstract: The molecular basis of beta-thalassemia is predominantly point mutations in the beta-globin gene. Frameshift 41-42 (-CTTT), IVS-2 position 654 (C-->T) mutation, nonsense codon 17 (A-->T), TATA box position -28 (A-->G) mutation and frameshift 71-72 (+A) account for more than 95% of beta-thalassemia alleles in the population of South China. We have developed a polymerase chain reaction (PCR)-mediated restriction fragment length polymorphism (RFLP) method for the identification of these alleles. In this method, artificial mispairing bases in PCR-amplified products were created to distinguish normal from mutant alleles on the basis of RFLPs. The size of the five PCR-amplified DNA fragments that may potentially contain the above five types of mutations is 93 or 89 bp (codons 41-42), 221 bp (IVS-2 nt 654), 110 bp (codon 17), 123 bp (TATA box nt -28), and 97 or 98 bp (codons 71-72). After these fragments were digested with Hinc II, Mae III, Nhe I, EcoR I, and Dde I, respectively, the allele-specific RFLPs produced were analyzed by gel electrophoresis. DNA samples of 24 patients with the above five types of beta-thalassemia were investigated with the present method and allele-specific oligonucleotide (ASO) probing simultaneously. We used this method in the prenatal diagnosis of 14 Chinese families for beta-thalassemia. The results obtained by the present method correspond well with those by the ASO probe test.
authors.affiliation: Molecular Biology Laboratory, Nanfang Hospital, First Military Medical University, Guangzhou, People's Republic of China.
authors.forename: X M
authors.initials: XM
authors.lastname: Xu
authors.forename: W F
authors.initials: WF
authors.lastname: Ma
authors.forename: L L
authors.initials: LL
authors.lastname: Song
authors.forename: Q
authors.initials: Q
authors.lastname: Xu
authors.forename: J Z
authors.initials: JZ
authors.lastname: Zhang
mesh_terms: D000328:Adult; D000483:Alleles; D044466:Asian People / Q000235:genetics*; D001483:Base Sequence; D005190:Family; D005260:Female; D005315:Fetal Diseases / Q000235:genetics*; D005838:Genotype; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics*; D012150:Polymorphism, Restriction Fragment Length; D011247:Pregnancy; D011296:Prenatal Diagnosis; D016385:TATA Box; D017086:beta-Thalassemia / Q000235:genetics*