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Abstract
Artemisinin-based drugs are the most effective medicine for the malaria treatment. To date, the main method of artemisinin production is its extraction from wormwood plants Artemisia annua L. Due to the limitation of this source, considerable efforts are now directed to the development of methods for artemisinin production using heterologous expression systems. Artemisinin is a sesquiterpene lactone, synthesized through the cyclization of farnesyl diphosphate involved in other sesquiterpene biosynthetic systems. Chrysanthemum species as well as A. annua, belong to Asteraceae family, and had been characterized by containing highly content of sesquiterpenes and their precursors. This makes chrysanthemum a promising target for the production of artemisinin in heterologous host plants. Chrysanthemum (C. morifolium Ramat.) was transformed by Agrobacterium tumefaciens carrying with the binary vectors p1240 and p1250, bearing artemisinin biosynthesis genes coding: amorpha-4,11-diene synthase, artemisinic aldehyde Δ11(13) reductase, amorpha-4,11-diene monooxygenase (p1240 was targeted to the mitochondria and p1250 was targeted to the cytosol), cytochrome P450 reductase from A. annua, as well as yeast truncated 3-hydroxy-3-methylglutarylcoenzyme A reductase. This study obtained 8 kanamycin-resistant lines after transformation with the p1240 and 2 lines from p1250. All target genes were detected in 2 and 1 transgenic lines of the 2 vectors. The transformation frequency of all target genes were 0.33% and 0.17% for p1240 and p1250, relative to the total transformed explant numbers. RT-PCR analysis revealed the transcription of all transferred genes in two lines obtained after transformation with the p1240 vector, confirming the possibility of transferring genetic modules encoding entire biochemical pathways into the chrysanthemum genome. This holds promise for the development of a chrysanthemum-based expression system to produce non-protein substances, such as artemisinin.
Highlights
The most effective drugs for the treatment of malaria are drugs combined with artemisinin, the use of such drugs is recommended by WHO resolution (WHA60.18, May 2007)
The aim of this work was to explore the feasibility of agrobacterium-mediated transformation of chrysanthemum with artemisinin biosynthesis genes
Two variants of the vectors were used in our study—with localization of amorpha-4,11-diene known, especially among crop plant belonging to flavor herbs or medicines, such as chrysanthemum synthase in the cell cytoplasm (p1250)—or in mitochondria (p1240)
Summary
The most effective drugs for the treatment of malaria are drugs combined with artemisinin, the use of such drugs is recommended by WHO resolution (WHA60.18, May 2007).artemisinin and its derivatives are effective to a number of viruses, many lines of human tumor cells, some tropical parasitic diseases and a number of livestock diseases [1]. The most effective drugs for the treatment of malaria are drugs combined with artemisinin, the use of such drugs is recommended by WHO resolution (WHA60.18, May 2007). Artemisinin is a terpenic lactone found in wormwood plant (Artemisia annua L.). Plants 2020, 9, 537 demand for artemisinin-containing drugs [2,3]. Improvement of cultivation techniques and breeding of A. annua has not led to a significant improvement in the yield of artemisinin [4,5]. In this connection, the possibility of producing artemisinin by methods alternative to extraction from natural sources is currently being actively studied. The biochemical pathway of artemisinin synthesis has been comprehensively studied [6,7] and this allowed advancing research on artemisinin production in heterologous systems
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