MOLECULAR DETECTION OF THE Cd 59 MUTATIONS IN THE HBA2 GENE AMONG POPULATIONS FORM SUMATERA, SULAWESI AND JAVA USING ACRS PCR-RFLP

Abstract

Thalassemia is a hereditary disorder with autosomal recessive inheritance pattern due to mutations in the coding of globin chains on hemoglobin formation. Mutations can occur in α-globin gene. In Indonesia, the number of people with thalassemia is increasing, and the number of carriers thalassemia were predicted 3-10% of the total population. World Health Organization launched a global strategy as a preventative and to control the number of people with thalassemia. One such strategy is the screening of carrier thalassemia. There are 834 people as research subjects, with the indication of carrier thalassemia were 61 individuals based on haematological screening thalassemia. This study aims to perform molecular detection of mutations non deletions, i.e Cd 59 mutation and Hb CS mutation by using Polymerase Chain Reaction (PCR) and PCR Restriction Fragment Length Polymorphisms (RFLP) with modifications primer using Artificially Created Restriction Site (ACRS) techniques against haematological results. Subject participants are archives DNA genomic thalassemia screening conducted by the Eijkman Institute for Molecular Biology in Sulawesi, Sumatra and Java population are not contained deletion mutations in two genes or one gene. Archival DNA genomic then amplified using specific primers to HBA2 gene sequence. Amplikon PCR results then analyzed by agarose gel RFLP method. Interpretation of the data was done by describing ribbon RFLP results by comparing the differences in migration distance of individual single stranded DNA of carrier thalassemia with normal individuals. Results showed one individual has a Cd 59 mutation on HBA2 gene with percentage of 0.53% means that there are 5 on 1000 people has Cd 59 mutations, this result is quite significant for Java population as a large population in Indonesia. PCR-RFLP was used to analyze genetic diversity among individuals within a population molecularly and as a method to detect heterozygosity and variations on the level of DNA sequences based on fragment length differences DNA caused by mutation.