PLASMODIUM FALCIPARUM RH5 INTERACTING PROTEIN (PFRIPR) FRAGMENT 811 – 1086 EXPRESSED IN ESCHERICHIA COLI BL21

• Main Author: Viki Zaneta, Michelia
• Format: Final Project
• Language: Indonesia
• Subjects: Kimia
• Online Access: https://digilib.itb.ac.id/gdl/view/87966
• Institution: Institut Teknologi Bandung

Malaria is one of the endemic diseases in Indonesia transmitted by female Anopheles mosquitoes, with significant prevalence in several regions such as 12,909 cases in East Nusa Tenggara Province, 216,380 cases in Papua Province, 7,079 cases in West Papua Province, and several areas in Kalimantan and Sumatra that remain highly endemic. Malaria can be prevented, one of which is through vaccination. One approach to developing an effective malaria vaccine is by forming a subunit vaccine using essential antigens in parasite growth to produce recombinant proteins. One promising antigen candidate is Plasmodium falciparum reticulocyte homologue 5 (Rh5) interacting protein (PfRIPR). This study aims to express the PfRIPR gene in E. coli BL21 (DE3) as a malaria vaccine candidate. The existence of PfRIPR protein has been proven to play a role in successful invasion so that antibodies against PfRIPR will inhibit merozoites in invading erythrocytes, thus hampering the growth of Plasmodium. The stages of this research include (i) amplification of the PfRIPR gene fragment AA 811 1086 with P. falciparum Jayapura isolate gDNA as a DNA template, (ii) construction of recombinant plasmid pET-30a-PfRIPR, (iii) transformation of E. coli BL21 (DE3) and E. coli BL21 (DE3) Codon Plus RIPL with recombinant plasmid pET-30a-PfRIPR through the heat shock method, (iv) expression of the PfRIPR gene fragment AA 811–1086 in E. coli BL21 (DE3) and E. coli BL21 (DE3) Codon Plus RIPL, (v) analysis of gene expression results using SDS-PAGE and Western Blot. The PfRIPR gene fragment AA 811–1086 was successfully amplified at an annealing temperature of 54.5°. The results of colony PCR, restriction analysis, and sequencing of the cloning product, the recombinant plasmids pGEM-T-PfRIPR and pET 30a-PfRIPR were successfully constructed. The recombinant plasmid pET-30a-PfRIPR fragment 811-1086 was successfully transformed into E. coli BL21 (DE3) and E. coli BL21 (DE3) Codon Plus RIPL. SDS-PAGE analysis showed that the recombinant PfRIPR protein fragment 811-1086, approximately 37 kDa in size, was not successfully expressed in E. coli BL21 (DE3) and E. coli BL21 (DE3) Codon Plus RIPL.