Expression of Two Key Enzymes of Artemisinin Biosynthesis FPS and ADS Genes in

Overview

Journal Adv Pharm Bull

Date 2021 Mar 22

PMID 33747865

Authors

Elfahmi

Rizqiya Astri Hapsari

Tamara Chrysanthy

Vaniarta Synthiarini

Fifi Fitriyah Masduki

Agus Setiawan

Toshiya Muranaka

Affiliations

Soon will be listed here.

Abstract

Artemisinin, a secondary metabolite in Artemisia annua is one of primary choice for the treatment of malaria, it is naturally produced in low concentration from this plant. This study was aimed to clone key enzymes of artemisinin production in order to enhance its production through the semi-synthetically production in Two key enzymes in artemisinin biosynthetic pathway which are farnesyl phosphate synthase () and amorpha-4,11-diene synthase (ads) genes were transformed into using pBEVY vector. Successful transformation was checked by polymerase chain reaction (PCR) method and sequencing analysis Recombinant plasmids which are pBEVY-GU_ and pBEVY_GL_ were successfully constructed. The optimized gene was amplified using PCR with a couple of primers that are designed in order to provide the homolog recombination between gene with the expression plasmid of pBEVY-GU respectively. While the optimized gene was cloned using classical method. Transformants were grown in selective media Synthetic Defined (SD) without leucine for transformants contain plasmid pBEVY-GL_ and media without uracil for transformants contain plasmid pBEVY-GU_. Confirmation of colonies was done by PCR with primers to amplify and . DNA from yeast was isolated from positive colonies then transformed to . Plasmid from was isolated for restriction analysis and sequencing. Protein expression was induced by cultivating the yeast in the media with 2% galactose. Based on PCR, restriction and sequencing analysis, it could be concluded that and genes were successfully constructed, transformed and expressed in S. cerevisiae.

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