KLONING GEN PENGKODE ENZIM FARNESYL PYROPHOSPHATE SYNTHASE (FPS) DARI TANAMAN ARTEMISIA ANNUA L.

• Main Author: Arifa Arbuati, Arrie
• Format: Final Project
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/45492
• Institution: Institut Teknologi Bandung

Artemisinin is a secondary metabolite produced by Artemisia annua plant, which is used as antimalarial drugs as Artemisinin-based Combination Therapy (ACTs). Artemisinin production in A. annua is relatively low, therefore efforts to enhance its production has been done using biotechnology and tissue culture methods. This method is focusing on the enzymes that have roles in artemisinin biosynthetic process in A. annua.When the enzymes could be overexpressed then influence the enhancement of the artemisinin production. One of the enzymes that has roles in artemisinin biosynthetic is farnesyl pyrophosphat synthase (FPS). This riset aimed to clone FPS gene in E.coli that can be furthermore transformed to A. annua plants. To obtain FPS gene clones, genomic DNA was isolated from A. annua leaves. Isolated genomic DNA was amplified by PCR using spesific primers of FPS gene. PCR product was checked using electrophoresis. Electroforegram result showed band at size around 1000 bp, fitting with FPS gene size which is 1032 bp. PCR product was ligated with cloning vectors PGEM-T Easy, and transformed into Escherichia coli. Plasmids in grown white colonies was isolated. Cloned confirmation was done with PCR amplification using FPS gene specific primers, enzyme restriction analysis using BglII enzyme which disgested FPS gene, EcoRI enzyme which disgested FPS gene out of plasmid, and sequencing of DNA nucleotide sequence. Confirmation results using PCR amplification and restriction analysis indicated to FPS gene, but the results of sequencing analysis showed the gene that had been inserted into the pGEM-T Easy vector was actually not the FPS gene. Therefore, the cloning of FPS enzyme gene to Escherichia coli using pGEM-T Easy was not succesful yet.