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《Journal of Plant Interactions》2013,8(1):811-824
Artemisinin is frequently used in the artemisinin-based combination therapy to cure drug-resistant malaria in Asian subcontinent and large swath of Africa. The hairy root system, using the Agrobacterium rhizogenes LBA 9402 strain to enhance the production of artemisinin in Artemisia annua L., is developed in our laboratory. The transgenic nature of hairy root lines and the copy number of transgene (rol B) were confirmed using polymerase chain reaction and Southern Blot analyses, respectively. The effect of different concentrations of methyl jasmonate (MeJA), fungal elicitors (Alternaria alternate, Curvularia limata, Fusarium solani, and Piriformospora indica), farnesyl pyrophosphate, and miconazole on artemisinin production in hairy root cultures were evaluated. Among all the factors used individually for their effect on artemisinin production in hairy root culture system, the maximum enhancement was achieved with P. indica (1.97 times). Increment of 2.44 times in artemisinin concentration by this system was, however, obtained by combined addition of MeJA and cell homogenate of P. indica in the culture medium. The effects of these factors on artemisinin production were positively correlated with regulatory genes of MVA, MEP, and artemisinin biosynthetic pathways, viz. hmgr, ads, cyp71av1, aldh1, dxs, dxr, and dbr2 in hairy root cultures of A. annua L. 相似文献
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Artemisinin, the endoperoxide sesquiterpene lactone, is an effective antimalarial drug isolated from the Chinese medicinal plant Artemisia annua L. Due to its effectiveness against multi-drug-resistant cerebral malaria, it becomes the essential components of the artemisinin-based combination therapies which are recommended by the World Health Organization as the preferred choice for malaria tropica treatments. To date, plant A. annua is still the main commercial source of artemisinin. Although semi-synthesis of artemisinin via artemisinic acid in yeast is feasible at present, another promising approach to reduce the price of artemisinin is using plant metabolic engineering to obtain a higher content of artemisinin in transgenic plants. In the past years, an Agrobacterium-mediated transformation system of A. annua has been established by which a number of genes related to artemisinin biosynthesis have been successfully transferred into A. annua plants. In this review, the progress on increasing artemisinin content in A. annua by transgenic approach and its future prospect are summarized and discussed. 相似文献
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Annual wormwood (Artemisia annua L.) produces an array of complex terpenoids including artemisinin, a compound of current interest in the treatment of drug-resistant
malaria. However, this promising antimalarial compound remains expensive and is hardly available on the global scale. Synthesis
of artemisinin has not been proved to be feasible commercially. Therefore, increase in yield of naturally occurring artemisinin
is an important area of investigation. The effects of inoculation by two arbuscular mycorrhizal (AM) fungi, Glomus macrocarpum and Glomus fasciculatum, either alone or supplemented with P-fertilizer, on artemisinin concentration in A. annua were studied. The concentration of artemisinin was determined by reverse-phase high-performance liquid chromatography with
UV detection. The two fungi significantly increased concentration of artemisinin in the herb. Although there was significant
increase in concentration of artemisinin in nonmycorrhizal P-fertilized plants as compared to control, the extent of the increase
was less compared to mycorrhizal plants grown with or without P-fertilization. This suggests that the increase in artemisinin
concentration may not be entirely attributed to enhanced P-nutrition and improved growth. A strong positive linear correlation
was observed between glandular trichome density on leaves and artemisinin concentration. Mycorrhizal plants possessed higher
foliar glandular trichome (site for artemisinin biosynthesis and sequestration) density compared to nonmycorrhizal plants.
Glandular trichome density was not influenced by P-fertilizer application. The study suggests a potential role of AM fungi
in improving the concentration of artemisinin in A. annua. 相似文献
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Artemisinin, a sesquiterpene lactone isolated from the Chinese medicinal plant Artemisia annua L., is an effective antimalarial agent, especially for multi-drug resistant and cerebral malaria. To date, A. annua is still the only commercial source of artemisinin. The low concentration of artemisinin in A. annua, ranging from 0.01 to 0.8% of the plant dry weight, makes artemisinin relatively expensive and difficult to meet the demand
of over 100 million courses of artemisinin-based combinational therapies per year. Since the chemical synthesis of artemisinin
is not commercially feasible at present, another promising approach to reduce the price of artemisinin-based antimalarial
drugs is metabolic engineering of the plant to obtain a higher content of artemisinin in transgenic plants. In the past decade,
we have established an Agrobacterium-mediated transformation system of A. annua, and have successfully transferred a number of genes related to artemisinin biosynthesis into the plant. The various aspects
of these efforts are discussed in this review. 相似文献
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Moran Farhi Magali Kozin Shai Duchin 《Biotechnology & genetic engineering reviews》2013,29(2):135-148
Artemisinin, a natural compound from Artemisia annua, is highly effective in treating drug-resistant malaria. Because chemical synthesis of this natural terpenoid is not economically feasible, its only source remains as the native plant which produces only small quantities of it, resulting in a supply that is far short of demand. Extensive efforts have been invested in metabolic engineering for the biosynthesis of artemisinin precursors in microbes. However, the production of artemisinin itself has only been achieved in plants. Since, A. annua possesses only poorly developed genetic resources for traditional breeders, molecular breeding is the best alternative. In this review, we describe the efforts taken to enhance artemisinin production in A. annua via transgenesis and advocate metabolic engineering of the complete functional artemisinin metabolic pathway in heterologous plants. In both cases, we emphasize the need to apply state-of-the-art synthetic biology approaches to ensure successful biosynthesis of the drug. 相似文献
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Pamela J. Weathers Shereen Elkholy Kristin K. Wobbe 《In vitro cellular & developmental biology. Plant》2006,42(4):309-317
Summary Artemisinin is a sesquiterpene lactone isolated from the aerial parts of Artemisia annua L. plants. Besides being currently the best therapeutic against both drug-resistant and cerebral malaria-causing strains
of Plasmodium falciparum, the drug has also been shown to be effective against other infections diseases including schistosomiasis and hepatitis.
More recently, it has also been shown to be effective against numerous types of tumors. Although chemical synthesis of artemisinin
is possible, it is not economically feasible. The relatively low yield (0.01–0.8%) of artemisinin in A. annua is a further serious limitation to the commercialization of the drug. Therffore, the enhanced production of artemisinin either
in cell/tissue culture or in the whole plant of A. annua is highly desirable. A better understanding of the biochemical pathway leading to the synthesis of artemisinin and its regulation
by both exogenous and endogenous factors is essential for facilitating increased yield. Two genes of the artemisinin biosynthetic
pathway have now been identified. This critical review covers recent developments related to the biosynthesis of this important
compound and related terpenoids, their regulation, and the production of these compounds both in vitro and in whole plants. 相似文献
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Characterization of development and artemisinin biosynthesis in self-pollinated <Emphasis Type="Italic">Artemisia annua</Emphasis> plants 总被引:1,自引:0,他引:1
Artemisia annua L. is the only natural resource that produces artemisinin (Qinghaosu), an endoperoxide sesquiterpene lactone used in the
artemisinin-combination therapy of malaria. The cross-hybridization properties of A. annua do not favor studying artemisinin biosynthesis. To overcome this problem, in this study, we report on selection of self-pollinated
A. annua plants and characterize their development and artemisinin biosynthesis. Self-pollinated F2 plants selected were grown under
optimized growth conditions, consisting of long day (16 h of light) and short day (9 h of light) exposures in a phytotron.
The life cycles of these plants were approximately 3 months long, and final heights of 30–35 cm were achieved. The leaves
on the main stems exhibited obvious morphological changes, from indented single leaves to odd, pinnately compound leaves.
Leaves and flowers formed glandular and T-shaped trichomes on their surfaces. The glandular trichome densities increased from
the bottom to the top leaves. High performance liquid chromatography–mass spectrometry-based metabolic profiling analyses
showed that leaves, flowers, and young seedlings of F2 plants produced artemisinin. In leaves, the levels of artemisinin increased
from the bottom to the top of the plants, showing a positive correlation to the density increase of glandular trichomes. RT-PCR
analysis showed that progeny of self-pollinated plants expressed the amorpha-4, 11-diene synthase (ADS) and cytochrome P450
monooxygenase 71 AV1 (CYP71AV1) genes, which are involved in artemisinin biosynthesis in leaves and flowers. The use of self-pollinated
A. annua plants will be a valuable approach to the study of artemisinin biosynthesis. 相似文献
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The stacked over-expression of FPS, CYP71AV1 and CPR genes leads to the increase of artemisinin level in Artemisia annua L. 总被引:1,自引:0,他引:1
Yunfei Chen Qian Shen Yueyue Wang Tao Wang Shaoyan Wu Ling Zhang Xu Lu Fangyuan Zhang Weimin Jiang Bo Qiu Erdi Gao Xiaofen Sun Kexuan Tang 《Plant biotechnology reports》2013,7(3):287-295
Artemisinin is an endoperoxide sesquiterpene lactone isolated from the aerial parts of Artemisia annua L., and is presently the most potent anti-malarial drug. Owing to the low yield of artemisinin from A. annua as well as the widespread application of artemisinin-based combination therapy recommended by the World Health Organization, the global demand for artemisinin is substantially increasing and is therefore rendering artemisinin in short supply. An economical way to increase artemisinin production is to increase the content of artemisinin in A. annua. In this study, three key genes in the artemisinin biosynthesis pathway, encoding farnesyl diphosphate synthase, amorpha-4, 11-diene C-12 oxidase and its redox partner cytochrome P450 reductase, were over-expressed in A. annua through Agrobacterium-mediated transformation. The transgenic lines were confirmed by Southern blotting and the over-expressions of the genes were demonstrated by real-time PCR assays. The HPLC analysis showed that the artemisinin contents in transgenic lines were increased significantly, with the highest one found to be 3.6-fold higher (2.9 mg/g FW) than that of the control. These results demonstrate that multigene engineering is an effective way to enhance artemisinin content in A. annua. 相似文献
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Artemisinin isolated from the aerial parts of Artemisia annua L. is a promising and potent antimalarial drug which has a remarkable activity against chloroquine-resistant and chloroquine-sensitive strains of Plasmodium falciparum, and is useful in treatment of cerebral malaria. Because the low content (0.01–1 %) of artemisinin in A. annua is a limitation to the commercial production of the drug, many research groups have been focusing their researches on enhancing the production of artemisinin in tissue culture or in the whole plant of A. annua. This review mainly focuses on the progresses made in the production of artemisinin from A. annua by biotechnological strategies including in vitro tissue culture, metabolic regulation of artemisinin biosynthesis, genetic engineering, and bioreactor technology. 相似文献
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Gao-Jie Hong Wen-Li Hu Jian-Xu Li Xiao-Ya Chen Ling-Jian Wang 《Plant Molecular Biology Reporter》2009,27(3):334-341
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Pamela J. Weathers Patrick R. Arsenault Patrick S. Covello Anthony McMickle Keat H. Teoh Darwin W. Reed 《Phytochemistry Reviews》2011,10(2):173-183
Artemisia annua L. produces the sesquiterpene lactone, artemisinin, a potent antimalarial drug that is also effective in treating other parasitic
diseases, some viral infections and various neoplasms. Artemisinin is also an allelopathic herbicide that can inhibit the
growth of other plants. Unfortunately, the compound is in short supply and thus, studies on its production in the plant are
of interest as are low cost methods for drug delivery. Here we review our recent studies on artemisinin production in A. annua during development of the plant as it moves from the vegetative to reproductive stage (flower budding and full flower formation),
in response to sugars, and in concert with the production of the ROS, hydrogen peroxide. We also provide new data from animal
experiments that measured the potential of using the dried plant directly as a therapeutic. Together these results provide
a synopsis of a more global view of regulation of artemisinin biosynthesis in A. annua than previously available. We further suggest an alternative low cost method of drug delivery to treat malaria and other
neglected tropical diseases. 相似文献
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1-Deoxy-d-xylulose-5-phosphate reductoisomerase (DXR), an important enzyme in the 2-c-methyl-d-erythritol-4-phosphate (MEP) pathway in plant plastids, provides the basic five-carbon units for isoprenoid biosynthesis. To investigate the roles of the MEP pathway in regulating growth, development and artemisinin biosynthesis of Artemisia annua L., we used RNA interference technology to generate transgenic plants with suppressed expression of DXR in A. annua (AaDXR). Suppression of AaDXR resulted in shorter stems, decreased branch numbers and leaf area, lower density of leaf trichomes. Although AaDXR-RNAi plants had no significant changes on the stomatal conductance, the net photosynthesis rate was decreased by 20.0–31.4% due to the marked decline in the contents of chlorophyll. Decreased levels of endogenous gibberellic acid (GA3) and abscisic acid were also detected in the transgenic lines. The artemisinin contents in leaves of all tested transgenic lines declined by 41.8–73.4% at the vegetative stage and 61.5–63.6% at the stages of flowering. The enhancement of artemisinin contents by methyl jasmonate at 300 µM has been abolished at seedling and vegetative stages in AaDXR-RNAi plants. These results demonstrate that AaDXR play import roles in the control of plan vegetative growth and artemisinin biosynthesis in A. annua. 相似文献
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Pu Shi Xueqing Fu Meng Liu Qian Shen Weimin Jiang Ling Li Xiaofen Sun Kexuan Tang 《Plant Cell, Tissue and Organ Culture》2017,129(2):251-259
Artemisinin, isolated from an annual herbaceous plant Artemisia annua L., is an effective antimalarial compound. However, artemisinin is accumulated in small amounts (0.01–0.1% leaf dry weight) in A. annua, resulting in constant high artemisinin price. Although metabolic engineering of partial artemisinin metabolic pathway in yeast achieved great success, artemisinin from A. annua is still the important business resource. Here, we report on the generation of transgenic plants with simultaneously overexpressing four artemisinin biosynthetic pathway genes, amorpha-4,11-diene synthase gene (ADS), amorpha-4,11-diene 12-monooxygenase gene (CYP71AV1), cytochrome P450 reductase gene (CPR), and aldehyde dehydrogenase 1 gene (ALDH1) via Agrobacterium-mediated transformation. The qRT-PCR analysis demonstrated that the introduced four genes of the transgenic lines were all highly expressed. Through high-performance liquid chromatography analysis, the artemisinin contents were increased markedly in transformants, with the highest being 3.4-fold higher compared with non-converter. These results indicate that overexpression of multiple artemisinin biosynthetic pathway genes is a promising approach to improve artemisinin yield in A. annua. 相似文献
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Chenfei Ma Huahong Wang Xin Lu Hong Wang Guowang Xu Benye Liu 《Metabolomics : Official journal of the Metabolomic Society》2009,5(4):497-506
Amorpha-4,11-diene synthase (ADS) is a very important enzyme which catalyzes the committed step of artemisinin biosynthesis.
In this work, two lines of transgenic Artemisia annua L. which ADS was over-expressed (line A9) and suppressed (line Amsi), respectively, were utilized. And the transgenic line
GUS with β-Glucuronidase gene was regarded as the control. Their terpenoid metabolic profiling was investigated by using GC × GC–TOFMS.
The metabolic profiling method established included simple extraction, two-dimension separation and multivariate analysis.
Partial least squares discriminant analysis (PLS-DA) was used to classify two transgenic lines and the control line. Eleven
important compounds in classification were identified. Most of them were sesquiterpenoids including monoterpenoid, diterpenoid
and four bioprecursors of artemsisnin. Compared with the control, artemisinin and bioprecursors in the line A9 increased as
a result of over-expressing ADS. Borneol and phytol also increased in the line A9, but (E)-β-farnesene and germacrene D were
reversely altered. The result indicated that over-expression of the ADS affected not only artemisinin biosynthesis, but also
the whole metabolic network of terpenoid. Compared with the line A9, no opposite change of artemisinin and related derivatives
was observed in the line Amsi, the ADS inhibition had no significant effect on artemisinin biosynthesis in the line Amsi. 相似文献
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Ling Zhang Xu Lu Qian Shen Yunfei Chen Tao Wang Fangyuan Zhang Shaoyan Wu Weimin Jiang Pin Liu Lida Zhang Yueyue Wang Kexuan Tang 《Plant Molecular Biology Reporter》2012,30(4):838-847
Artemisinin has attracted interest due to its medicinal value in treating malaria and its potential for use against certain cancers and viral diseases. Trichome density and capacity determine artemisinin content in Artemisia annua plants. Thus, the ATP-binding cassette transporter G (ABCG) subfamily involved in trichome cuticle development may also influence artemisinin accumulation. In this study, putative A. annua ABC transporter unigenes were identified and classified from the unigene sequences up to date in the National Center for Biotechnology Information database, and nine putative A. annua ABCG transporter unigenes that may be involved in cuticle development were selected for expression analyses. Two of them, AaABCG6 and AaABCG7, showed parallel expression pattern as two artemisinin biosynthesis-specific genes (amorpha-4, 11-diene synthase and a cytochrome P450-dependent hydroxylase, CYP71AV1) in different tissues and different leaf development stages and also showed similar induction in the plants after methyl jasmonate or abscisic acid treatments. Identification of these putative A. annua ABCG transporter unigenes could provide the basis for cloning of the full-length genes and further functional investigation to find the artemisinin relevant transporters, which could be used for improving artemisinin yield in both A. annua plants and heterologous systems using transgenic technology. 相似文献
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Artemisinin is a promising and potent antimalarial drug naturally produced by the plant Artemisia annua L. but in very low yield. Its artemisinin content is known to be greatly affected by both genotype and environmental factors.
In this study, the production of artemisinin and leaf biomass in Artemisia annua L. was significantly increased by exogenous GA3 treatment. The effect of GA3 application on expression of proposed key enzymes involved in artemisinin yield was examined in both wild type (007) and
FPS-overexpression (253-2) lines of A. annua. In the wild type (007) at 6 h post GA3 application there was an abrupt rise in FPS, ADS and CYP71AV1 expression and at 24 h a temporary and significant peak in artemisinin (1.45-fold higher than the control). After GA3 application in line 253-2, there was a dramatic rise in expression of FPS at 3 h, CYP71AV1 at 9 h and ADS at 72 h and accumulation of artemisinin after 7 days, which was a delay when compared with the wild type plant. Thus, increased
artemisinin content from exogenous GA3 treatment was associated with increased expression of key enzymes in the artemisinin biosynthesis pathway. Interestingly,
exogenous GA3 continuously enhanced artemisinin content from the vegetative stage to flower initiation in both plant lines and gave significantly
higher leaf biomass than in control plants. Consequently, the artemisinin yield in GA3-treated plants was much higher than in control plants. Although the maximum artemisinin content was found at the full blooming
stage [2.1% dry weight (DW) in 007 and 2.4% DW in 253-2], the highest artemisinin yield in GA3-treated plants was obtained during the flower initiation stage (2.4 mg/plant in 007 and 2.3 mg/plant in 235-2). This was
26.3 and 27.8% higher, respectively, than in non-treated plants 007 and 253-2. This study showed that exogenous GA3 treatment enhanced artemisinin production in pot experiments and should be suitable for field application. 相似文献