Detection of beta-globin gene mutations among Kelantan Malay thalassaemia patients by polymerase chain reaction restriction fragment length polymorphism.

INTRODUCTION: Beta-thalassaemia major is an autosomal recessive disorder that results in severe microcytic, hypochromic, haemolytic anaemia among affected patients. Beta-thalassaemia has emerged as one of the most common public health problems in Malaysia, particularly among Malaysian Chinese and Malays. This study aimed to observe the spectrum of mutations found in Kelantan Malay beta-thalassaemia major patients who attended the Paediatrics Daycare Unit, Hospital Universiti Sains Malaysia, Kelantan, Malaysia, the data of which was being used in establishing the prenatal diagnosis in this Human Genome Centre. METHODS: This was a cross-sectional study conducted with 35 Kelantan Malay beta-thalassaemia major patients. DNA was extracted from the blood collected from the patients and subjected to polymerase chain reaction (PCR) amplification. Six restriction enzymes were used to digest the PCR products for the detection of mutations. RESULTS: Five out of the six beta-globin gene defects were detected, namely, IVS-1 nt5 (G>C), IVS-1 nt1 (G>T), codon 26 (G>A), codon 41-42 (4 bp del) and codon 19 (A>G). The mutation which was not observed in this study was in codon 15 (G>A). The two most common mutations observed were codon 26 (G>A) and IVS-1 nt5 (G>C), which was detected in 26 and 17 patients, respectively. Two patients did not show any of the six mutations. CONCLUSION: Our results added to the existing data on the common beta-globin gene defects in Kelantan Malay beta-thalassaemia patients.

An improved rapid genotyping method for the study of beta-2 adrenergic receptor gene polymorphisms using single tube allele specific multiplex PCR.

BACKGROUND: Seventeen single nucleotide polymorphisms (SNPs) have been identified so far, within the beta-2 receptor (beta(2) AR) gene. The presence of so many SNPs within the beta(2) AR gene causes a problem, for those studying beta(2) AR pharmacogenetics, in relation to which SNPs to choose. Most of the work has focused on the three common SNPs within the coding block (alleles 16, 27 and 164) and the techniques developed have been for these three functionally important alleles. OBJECTIVE: We report an improved polymerase chain reaction (PCR)-based method for the simultaneous detection of five functionally important beta(2) AR alleles, namely beta 16A/G, beta utr-20C/T, beta 27C/G, beta utr-47C/T and beta 164C/T. METHODS: Genomic DNA was used as a template for duplex and triplex PCR to detect the polymorphic sites of the five alleles. RESULT: DNA sequencing analysis confirmed the specificity of this PCR method. CONCLUSION: This simplified single-tube multiplexed PCR assay provides an easier, faster and more cost-effective method than those available for studying the specified polymorphisms of the beta(2)AR gene.