CLONING OF PLASMODIUM FALCIPARUM ERITHROCYTE BINDING ANTIGEN 175 (PFEBA-175) RIII-V GENE FROM TIMIKA ISOLATE FOR MALARIA VACCINES DEVELOPMENT

• Main Author: Aditya Nugraha, Dadi
• Format: Final Project
• Language: Indonesia
• Subjects: Kimia
• Online Access: https://digilib.itb.ac.id/gdl/view/35736
• Institution: Institut Teknologi Bandung

175 kDa Plasmodium falciparum erythrocyte binding antigen (PfEBA-175) is a protein that plays an important role in human erithrocyte cell invasion by a parasite, Plasmodium falciparum, which cause malaria disease. This protein is a potential malaria vaccine candidate as it can induces antibody production in human body which inhibit malaria parasite invasion. PfEBA-175 has been known to binding glycoporin A (GpA), a receptor protein in erithrocyte cell membrane surface during invasion. PfEBA-175 shares similar form with type I transmembrane protein and it’s extracelular has 6 regions. Antibodies for RIII-V has been shown can inhibit malaria parasite growth and protect patient from clinical malaria symptoms. In this study, gene that encodes PfEBA-175 RIII-V (ef3-5) has been cloned by standard methods. The gene was amplified by PCR using spesific primers (Ta=63oC, increment -2 oC /cycle) and genomic DNA of local isolate P. falciparum was used as a template. The amplicons were inserted to cloning vector, pGEM-t Easy through ligation to get recombinant plasmids, pGEMef3-5. The recombinant plasmid was transformed into E. coli TOP10 F’ for maintaining. The pGEMef3-5 was extracted from transformant E. coli and after that, the plasmid were evaluated by restriction digestion analysis using enzymes and were confirmed by DNA. DNA alignment of pGEMef3-5 and ef3-5 (3D7) shows that the cloning was performed correctly with similarity, 99.63% (clone 4) and 99.50% (clone 10). DNA sequencing analysis shows clone 4 have base mutation at base position, 23(G?A), 542 (T?C), 939 and 1313 (A?G) and 1019 (G?A). Clone 10 have base mutation at base position, 23 (G?A), 542 (T?C), 939 and 1313 (A?G) and deletion at 458 and 460. The Results indicate this mutations might be genetic variation resulted from isolate