Ficus carica hairy roots: In vitro anti-leishmanial activity against Leishmania major promastigotes and amastigotes

Objective: To investigate the biochemical capacity, and in vitro inhibitory effects of hairy roots from two cultivars of Ficus carica L. (Sabz and Siah) on Leishmania major promastigotes and amastigotes. Methods: In the hairy roots, the activity of antioxidant enzymes compared to normal leaves and roots, and the presence of some phenolic compounds in comparison with fruits were investigated. The IC 50 values of hairy roots in promastigotes was determined by tetrazolium-dye 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide and trypan blue assays. By calculating the infectivity index of peripheral blood mononuclear cells (PBMCs), the leishmanicidal activity (IC 50 values) of hairy roots for amastigotes was estimated. The effects of hairy roots (IC 50 values) treatment on the levels of IFN-γ and iNOS expression, intracellular reactive oxygen species, and iNOS protein expression in infected-PBMCs were determined. Results: Based on antioxidant enzyme assays and high performance liquid chromatography analysis, hairy roots exhibited high antioxidant capacity and contained high levels of phenolic compounds. According to the results of tetrazolium-dye 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide and trypan blue assays, the hairy root extracts of both cultivars showed considerable dose-dependent inhibitory effects against Leishmania major promastigotes. Depending on the concentration and exposure time, treatment of infected-PBMCs with hairy root extracts caused the generation of a significant reactive oxygen species, up-regulation of IFN-γ and iNOS genes expression, and high value of iNOS protein compared to controls. Conclusions: The findings of this study suggest that the hairy roots of Ficus carica can be considered as a promising natural source of antileishmanial agents.

First Report of Agrobacterium rhizogenes-induced Hairy Root Formation in Selaginella bryopteris: a Pteridophyte Recalcitrant to Genetic Transformation

Abstract we report A. rhizogenes-induced hairy root formation in S. bryopteris, a medicinally and commercially important plant. A. rhizogenes strain LBA1334 co-cultivated with explants (root, rhizophore, stem portion near the root, and stem with intact fronds) for 24 and 48 h after transformation for induction of hairy roots. The induction of hairy root was observed after 6 days of infection in case of 48 h co-cultivation only. PCR with rolA and virC gene specific primers confirmed the induced hairy roots were due to Ri T-DNA integration and not due to contaminating A. rhizogenes. The root network as explants showed the maximum transformation efficiency. We tested different media like MS, SHFR (Stage Hog Fern Root) and KNOP's during transformation for hairy root induction. The SHFR based media showed good response in transformation as well as propagation. Further, transformation efficiency was enhanced by addition of TDZ (2 mg/L) and Bevistin (0.1%) in SHFR media. The present work would be helpful in hairy roots-based in vitro production of secondary metabolites and on aspect of functional genomics of S. bryopteris.

Effects of different nitrogen sources on accumulation of active components in hairy roots of Salvia miltiorrhiza and Salvia castanea f. tomentosa / 中草药

Objective: Nitrogen is an important element affecting the accumulation of effective components in Chinese medicinal materials. The purpose of this study was to investigate effects of different nitrogen sources on the growth and active components accumulation of hairy roots of Salvia miltiorrhiza and Salvia castanea f. tomentosa. Methods: The hairy roots of S. miltiorrhiza and S. castanea f. tomentosa were treated with ammonium nitrate, hydrolyzed milk protein, peptone, beef extract, casein and yeast extract, respectively. The growth of hairy roots and the accumulation of active components were analyzed. Results: Ammonium nitrate was the most beneficial to the growth of the two kinds of hairy roots. Hydrolyzed milk protein significantly promoted the accumulation of salvianolic acids, compared with ammonium nitrate, the contents of rosmarinic acid and salvianolic acid B in S. miltiorrhiza were respectively increased by 2.94 times and 3.27 times, and the contents of rosmarinic acid and salvianolic acid B in S. castanea f. tomentosa were respectively increased by 13.74 times and 2.01 times. Yeast extract had the most significant effect on the accumulation of dihydrotanshinone I and cryptotanshinone in two kinds of hairy roots. Hydrolyzed milk protein significantly promoted the accumulation of tanshinone IIA in the hairy roots of S. miltiorrhiza, while beef extract had the most significant effect on the accumulation of tanshinone IIA in S. castanea f. tomentosa. Conclusion: Ammonium nitrate was the best nitrogen source for the growth of two kinds of hairy roots, and hydrolyzed milk protein was the best nitrogen source for salvianolic acids accumulation. The effects of different nitrogen sources on four kinds of tanshinones were different, and the responses of S. miltiorrhiza and S. castanea f. tomentosa to different nitrogen sources were also different. This study not only has certain guiding significance for large-scale cultivation of hairy roots of S. miltiorrhiza and industrialized production of active components, but also provides a reference for the development and utilization of S. castanea f. tomentosa resources.

Effects of different carbon sources on growth and active component contents in Salvia miltiorrhiza and S. castanea f. tomentosa hairy roots / 中国中药杂志

Salvia miltiorrhiza(Sm) and Salvia castanea f. tomentosa(Sc) hairy roots were used as experimental materials to study the effects of six different carbon sources, galactose, fructose, lactose, glucose, arabinose and sucrose(control), on fresh weight, dry weight, contents and yields of salvianolic acids and tanshinones. The results showed that galactose was most beneficial to the growth of two kinds of hairy roots, while lactose and arabinose were not conducive to their growth. As for Sm hairy roots, fructose significantly promoted the accumulation of salvianolic acid B, and the content increased by 5.801 times and 10.151 times compared with the control group, respectively. Glucose significantly promoted the accumulation of salvianolic acids. The content and yield of rosmarinic acid were 7.674 times and 9.260 times of that of the control group, and the content and yield of salvianolic acid B were 5.532 times and 6.675 times of the control group. For the hairy roots of Sc, galactose significantly increased the content and yield of rosmarinic acid, reaching 7.820 times and 9.944 times of the control group, respectively. Fructose promoted the increase of the content and yield of cryptotanshinone, reaching 9.242 times and 6.609 times of the control group, respectively. The study confirmed the optimal carbon source for the hairy root culture of Sm and Sc, and provided theoretical guidance for large-scale production of Sm drug-derived components and the utilization of Sc.

Over-expressing root-specific β-amyrin synthase gene increases glycyrrhizic acid content in hairy roots of glycyrrhiza uralensis / 中草药·英文版

Objsective: Glycyrrhizia uralensis, one of the most widely-used traditional Chinese medicines, is mainly cropped in China. However, many cultivars are less in glycyrrhizic acid than Chinese Pharmacopoeia requires. In this paper, we improved glycyrrhizic acid by regulating β-amyrin synthase gene (GuBAS). Methods: Tobacco root-specific promoter TobRB7 and GuBAS cDNA were obtained and combined with linearized pCAMBIA1305.1 to construct root-specific plant expression vector which was later transformed into Agrobacterium rhizogenes ACCC10060 by electrotransformation. The cotyledons and hypocotyls of G. uralensis were infected by the recombinant A. rhizogenes ACCC10060 to induce hairy roots. The GA content was quantified by HPLC. Results: The PCR and sequencing results both showed that three transgenic hairy root lines were obtained. The copy number of GuBAS in these transgenic hairy roots was intended by qRT-PCR to be 3, 7, and 4. GA was detected by HPLC, and the results showed that GA was present in the three transgenic hairy roots, while absent in wild hairy roots. Conclusion: Over-expressing GuBAS root-specifically in hairy roots of G. uralensis enhanced GA accumulation.

Influence of elicitors on effective constituents and expression of key metabolic enzyme genes in hairy roots of Atropa belladonna / 中草药

Objective To improve the yield of the effective medicinal ingredients of Atropa belladonna, to provide economic and efficient method for the actual production of A. belladonna, therefore to provide a basic reference for the related research on the mechanism of secondary metabolism of medicinal plants. Methods In this study, the influences of four kinds of elicitors, including methyl jasmonate (MJ), AgNO3, salicylic acid (SA), yeast extract (YE), on the accumulation of active components and the expression level of key metabolic enzyme genes, including putrescine N-methyl transferase (pmt), tropinone reductase-I (trI), and hyoscyamine-6-β-hydroxylase (h6h), were studied in hairy roots of A. belladonna. 0.5 g fresh hairy roots of A. belladonna were cultivated in B5 liquid medium. 12 d later, these mediums were replaced with the new medium containing one kind of four elicitors. Hairy roots were taken samples after 2 d to mensurate the fresh weight, dry weight, the content of tropane alkaloids, some physiological indexes, and three key genes expression level. Results MJ inhibited the growth and tropane alkaloids accumulation of hairy roots. The gene expression level of pmt and trI also decreased compared with control group (CK). The contents of tropane alkaloids and the expression level of pmt, trI and h6h were all increased compared with CK by the treatment of AgNO3, while the growth of hairy rootswas inhibited; SA contributed to the increased content of hyoscyamine, but with no obvious influence on growth of hairy roots. As to YE, the content of tropane alkaloids and the expression level of pmt, trI, h6h were all increased correspondingly. Further more, YE was benefit for the growth of hairy roots. Conclusion Elicitors had selective influence on growth and tropane alkaloids accumulation in hairy roots of A. belladonna. The best elicitor on accumulation of tropane alkaloids was AgNO3. YE could effectively improve of the growth of hairy and contents of tropane alkaloids. This study concluded that these elicitors influence the secondary metabolism of A. belladonna by regulating and controlling the expression level of some genes of key metabolic Enzyme, which could provide an effective method to enhance the medicinal composition in the culture of hairy roots of A. belladonna.

Mechanism exploration on nitrogen metabolism and secondary metabolism in Atropa belladonna hairy roots treated with yeast extract / 中国中药杂志

In order to study the mechanism of nitrogen metabolism and secondary metabolism in Atropa belladonna hairy roots treated with yeast extract, yeast extract（YE） was added to the culture medium. Then the changes of physiological and biochemical indexes of A. belladonna hairy roots after treatment with YE were detected. The results are as follows，the activity of key enzymes of nitrogen metabolism changed differently. Compared with the control group (CK), the activity of nitrate reductase (NR) and glutamine synthetase (GS) were significantly increased, while the activity of glutamate dehydrogenase (GDH) was not changed significantly. The content of nitrate nitrogen, ammonium nitrogen had a significant decrease,but the content of soluble protein, free amino acid, total nitrogen are significantly more than CK. Moreover, YE treatment led to the increase of the content precursor amino acids (ornithine and arginine) and precursor putrescine in secondary metabolic pathways of A. belladonna. The expression level of gene putrescine N-methyl transferase (pmt), tropinone reductase-I (trI) and hyoscyamine 6-β-hydroxylase(h6h) all increased in a different rate caused by YE treatment, which eventually led to the increase of the yield of tropane alkaloids. The yield of hyoscyamine and scopolamine were 3.09 and 1.85 folds than that of CK after 16 days treatment time. The results indicated that YE can induce more synthesis of tropane alkaloids by increasing the activity of key enzymes in nitrogen metabolism to provide more synthetic materials for secondary metabolism, meanwhile it regulated the expression level of some genes of key metabolic enzyme to accelerate secondary metabolism.

Factors influencing induction and in vitro culture of hairy roots in Phytolacca americana L / 生物工程学报

To use hairy roots for producing medicinal ingredients of Phytolacca americana L. we studied the factors influencing the induction and in vitro culture. Hairy roots could be incited from the veins of cut surface (morphological lower) of P. americana L. leaf explants around 18 days after infection with the strain of Agrobacterium rhizogenes ATCC15834. The highest rooting rate, 70%, was obtained when leaf explants were pre-cultured for 1 day, infected for 20 min, and co-cultured for 4 days. The transformation was confirmed by PCR amplification of rolC of Ri plasmid and silica gel thin-layer chromatography of opines from P. americana L. hairy roots. All the hairy root lines could grow rapidly on solid exogenous phytohormone-free MS medium. Among the 9 hairy root lines, the hairy root line 2 had most rapid growth, most branched lateral roots and most intensive root hair; the root surface of some hairy root lines seemed purple or red, while that of the other hairy root line appeared white. Among liquid media MS, 1/2MS, B5 and 6,7-V tested, the best growth for hairy root lines was attained in liquid exogenous phytohormone-free MS medium. Compared with exogenous phytohormone-free MS medium, 6,7-V medium was better for synthesis and accumulation of esculento side A in hairy roots. The established optimal conditions for induction and in vitro culture of P. americana hairy roots had laid an experimental and technological foundation for production of medicinal constituents esculento side A from large scale culture of hairy roots.

Establishment of culture system of Vaccaria segetalis hairy roots and determination of vaccarin / 中草药

Objective: To establish the culture system of Vaccaria segetalis hairy roots. Methods: Agrobacterium rhizogenes R15834, R1601, R1000, and A4 were used to infect V. segetalis leaves to induce hairy roots and the influences of different physicochemical factors on its growth were investigated. The content of vaccarin was determined by HPLC. Results: A. rhizogenes R1601 had the highest induction rate by converted into V. segetalis leaves, The best growth cycle of cell suspension culture was defined when cultured in liquid MS medium with pH value of 6 and sucrose concentration of 3%, vaccarin in V. segetalis hairy roots was slightly higher than that in the seed. Conclusion: V. segetalis could successfully induce hairy roots, the foundation has been established for further oplimizing the proper cultural system for V. segetalis hairy roots and regulating the secondary metabolites.

Culture of hairy roots in Solanum lyratum and determination of diosgenin / 中草药

Objective: To establish the introduction and culture system of the hairy roots in Solanum lyratum and to screen the clone of hairy roots with more diosgenin. Methods: The explants of S. lyratum were infected by Agrobacterium tumefaciens strain C58C1, to obtain the hairy roots and construct the genetic transformation system of the hairy roots in S. lyratum. HPLC was used to determine the diosgenin in the hairy roots. Results: The optimum transformation results were obtained with the max inductivity of hairy roots of 83.33% during infecting time for 10 min by C58C1 and co-cultural time of 4 d. The average content of diosgenin in the hairy roots was 4.620 mg/g, it was 2.652 times as high as that in the leaves (1.742 mg/g) which had the highest diosgenin content in the different tissues of wild type plant of S. lyratum. Conclusion: It is an effective way to obtain diosgenin from the hairy roots of S. lyratum.

Study on content of terpenoid indole alkaloids and transcriptional differences for covalent orca3/g10h in double-gene transgenic hairy roots of Catharanthus roseus / 中草药

Objective: In order to improve the content of terpenoid indole alkaloids (TIAs), orca3/g10h genes were introduced to the hairy roots in Catharanthus roseus. Methods: Bivalent expression vector CAMBIA1304+ +orca3 + g10h was constructed and introduced into Agrobacterium rhizogenes strain and transformed into C. roseus to obtain transgenic hairy roots. RT-qPCR was used to study the transcriptional differences of relative genes referred to the biosynthesis pathway of TIAs. Then HPLC was used to study TIAs content in the transgenic hairy roots of C. roseus, including vinblastine, vincristine, and ajmalicine. Results: The transcriptional level of genes that linked to biosynthesis of TIAs in the transgenic hairy roots of C. roseus, asα, ggpps, g10h, str, tdc, cpr, sgd, and dat, were all expressed higher than those of the nontransgenic roots. HPLC results showed that modified hairy root of C. roseus owned more total TIAs production, 58.23 mg/g, the number was larger than that of common roots in C. roseus as many as 27.5 times. On the other hand, the average content of vinblastine and vincristine was also more than the common roots in C. roseus. Among them, vinblastine content was the most. The number of production got 51.30 mg/g, which was as many as 197.3 times of the common root of C. roseus. Conclusion: Orca3/g10h double-gene transgenic hairy root of C. roseus can increase TIAs content efficiently.

Effects of PEG-6000 stress on tanshinones accumulation in hairy roots of Salvia miltiorrhiza / 中草药

Objective: To study the effects of polyethylene glycol-6000 (PEG-6000) stress on the accumulation of tanshinones in hairy roots of Salvia miltiorrhiza. Methods: Agrobacterium rhizogenes ATCC15834 was used to induce the hairy roots of S. miltiorrhiza. After 20 d suspension culture, the PEG-6000 (1.2%, 2.0%, 5.5%, and 10%, respectively) was added into the suspension cultures and at the same time, the contents of tanshinones (including tanshinone I, cryptotanshinone, dihydrotanshinon I, and tanshinone IIA) were quantified by HPLC on day 7. Results: The growth of the hairy roots of S. miltiorrhiza was inhibited by PEG-6000. After PEG-6000 (1.2%, 2.0%, 5.5%, and 10%) treatment, the dry weights of the hairy roots of S. miltiorrhiza were reduced to 75.1%, 83.0%, 76.2%, and 76.1% of the control group, respectively. Addition of PEG-6000 at different levels could significantly increase the yields of four tanshinones in the hairy roots of S. miltiorrhiza. The yields of tanshinone I, cryptotanshinone, dihydrotanshinon I, and tanshinone IIA were significantly increased by 2.0%-5.5%, 1.2%, 2.0%, and 5.5% PEG-6000, respectively. And the tanshinone IIA increased most. Conclusion: PEG-6000 could stimulate the accumulation of tanshinones in the hairy roots of S. miltiorrhiza.

Carbohydrate and elicitor enhanced withanolide (withaferin A and withanolide A) accumulation in hairy root cultures of Withania somnifera (L.).

Leaves of Withania somnifera contained more withaferin A and withanolide A than roots indicating that these compounds mainly accumulate in leaves. With an increase in age of the plant, withaferin A was enhanced with a corresponding decrease in withanolide A. Hairy root cultures were induced from leaf explants using Agrobacterium rhizogenes and the transgenic nature of hairy roots was confirmed by partial isolation and sequencing of rolB gene, which could not be amplified in untransformed plant parts. In hairy roots, withaferin A accumulated at 2, 3 and 4% but not at 6% sucrose, the highest amount being 1733 mg/g dry weight at 4% level. High and equal amounts of withaferin A and withanolide A accumulated (890 and 886 mg/g dry tissue respectively) only at 3% sucrose. Increasing concentrations of glucose enhanced withaferin A and it peaked at 5% level (3866 mg/g dry tissue). This amount is 2842 and 34% higher compared to untransformed roots and leaves (collected from 210-day-old plants) respectively. Withanolide A was detected at 5% glucose but not at other concentrations. While chitosan and nitric oxide increased withaferin A, jasmonic acid decreased it. Acetyl salicylic acid stimulated accumulation of both withaferin A and withanolide A at higher concentrations. Triadimefon, a fungicide, enhanced withaferin A by 1626 and 3061% (not detected earlier) compared to hairy and intact roots respectively.

Efecto de la aplicación de elicitores sobre la producción de 4b-hidroxiwithanólido E, en raíces transformadas de Physalis peruviana L / Efeito da aplicação de elicitores sobre a produção de 4b- hidroxiwithanólido E, em raízes transformadas de Physalis peruviana L / Effect of elicitor application on the production of 4-b-hydroxy withanolide E by hairy roots of Physalis peruvianaEffect of elicitor application on the production of 4-b-hydroxy withanolide E by hairy roots of Physalis peruviana

Objetivo: Estudiar la producción del metabolito 4b-hidroxiwithanólido E, mediante el cultivo in vitro de raíces transformadas de uchuva (Physalis peruviana L.) y evaluar el efecto de la influencia de la aplicación de diferentes elicitores sobre la producción de dicho metabolito. Materiales y métodos: Se obtuvieron raíces transformadas de Physalis peruviana L. mediante infección con Agrobacterium rhizogenes C106. Se cultivaron las raíces transformadas en medio líquido Murashige & Skoog, durante cuatro semanas, al cabo de las cuales se aplicaron diferentes concentraciones de sulfato de cobre, ácido salicílico y ácido jasmónico durante 24 horas. Se cuantificó el contenido del metabolito por cromatografía líquida...

Effect of elicitor application on the production of 4-b-hydroxy withanolide E by hairy roots of Physalis peruviana. Objectives: To study the metabolite 4-b-hydroxy withanolide E production by the in vitro culture of golden berry (Physalis peruviana L.) transformed roots, and to evaluate the effect of different elicitors on the metabolite production. Materials and methods: Hairy roots of Physalis peruviana L were obtained through infection with Agrobacterium rhizogenes C106. Hairy roots were cultured on Murashige & Skoog liquid medium for four weeks, before being exposed to different concentrations of copper sulfate, salicylic acid and jasmonic acid during 24 hours. Metabolite contents were quantified using High Performance Liquid Chromatography. Results: The highest amount of 4-bhydroxy withanolide E in hairy root tissues (0.323 mg/g of dry roots) was obtained after exposing the tissues to 10 mM salicylic acid as elicitor. Conclusions: 4-b-hydroxy withanolide E production in hairy roots was improved by using elicitors such as salicylic acid and copper sulphate. The highest concentration of the metabolite in hairy roots treated with elicitors was 1.538 times the control concentration (without elicitor treatment)...

Efeito da aplicação de elicitores sobre a produção de 4b- hidroxiwithanólido E, em raízes transformadas de Physalis peruviana L. Objetivo: Estudar a produção do metabolito 4b-hidroxiwithanólido E em cultivo in vitro de raízes transformadas de “uchuva” (Physalis peruviana L.) e avaliar o efeito da influência da aplicação de diferentes elicitores na produção deste metabolito. Materiais e métodos: Foramobtidas raízes transformadas de Physalis peruviana L. através da infecção com Agrobacterium rhizogenes C106. Cultivaram-se durantequatro semanas as raízes transformadas em meio líquido Murashige & Skoog, depois se aplicaram diferentes concentrações de sulfato de cobre, ácido salicílico e ácido jasmónico durante 24 horas. Quantificou-se o conteúdo do metabolito por cromatografía líquida. Resultados: Ao quantificar a produção de 4b-hidroxiwithanólido E presente nos tecidos encontrou-se a maior produção (0,323mg/g raízes seca) após aplicação do ácido salicílico a uma concentração de 10 mM como elicitor. Conclusões: Aumentou-se a produção de 4b-hidroxiwithanólido E aplicando elicitores como o ácido salicílico e sulfato de cobre, nas raízes de Physalis peruviana L. transformadas com Agrobacteriumrhizogenes. A maior concentração do metabolito é 1,538 vezes a encontrada nas raízes não tratadas com elicitores...

Biotechnology approaches for production of antiulcerogenic dihydro-epideoxyarteannuin B isolated from Artemisia annua L. / Ferramentas biotecnológicas para a produção de dihidro-epideoxiarteanuina B, um antiulcerogênico isolado de Artemisia annua L.

Methodologies were developed for the establishment and cultivation of Artemisia annua L (CPQBA 2/39 x PL5 hybrid) roots submitted to light conditions and genetic transformation performed with Agrobacterium rhizogenes (15834 and 8196 strains). The transgenic and non-transgenic (normal) roots were cultured in Murashige and Skoog (1962) medium, kept under different photoperiodic conditions and analyzed for evaluation of the antiulcerogenic dihydro-epideoxyarteannuin B (compound A) contents. The Dot Blot technique was used to confirm the transgenic nature of the roots. The plants¢s crude extracts were analyzed by Gas Chromatography coupled to Mass Spectrum (CG/MS). The chromatograms of the extracts taken from normal roots revealed the presence of dihydro-epideoxyarteannuin B and other compound (compound B). Photoperiods during cultivation influenced the production of these two compounds: under continuous darkness dihydro-epideoxyarteannuin B was intensely produced and the compound B present in small amounts, while on 16 h photoperiod, the inverse occurred. The quantification of dihydro-epideoxyarteannuin B by Gas Chromatography coupled to Flame Detector Ionization (CG/FID) revealed an approximately fivefold increase in the production of this compound by normal roots kept under continuous darkness compared to roots kept under 16 h light period. The terpene dihydro-epideoxiarteannuin B was not present in transgenic hairy roots.

Foram desenvolvidas metodologias para o estabelecimento e cultivo de raízes de Artemisia annua L. (híbrido CPQBA 2/39 x PL5). Estas raízes foram submetidas a diferentes condições de luz e a transformação genética com Agrobacterium rhizogenes (cepas 8196 e 15834). As raízes transgênicas e não-transgênicas (normais) foram cultivadas em meios de Murashige e Skoog (1962), mantidas sobre diferentes condições de fotoperíodo e analisadas para avaliação do conteúdo do composto antiulcerogênico dehidro-epideoxiarteanuína B (composto A). A confirmação do caráter transgênico das raízes foi obtida por Dot Blot. Os extratos dos materiais vegetais foram analisados por Cromatografia Gasosa acoplada a um Espectômetro de Massas (CG/EM). Os cromatogramas dos extratos das raízes normais revelaram a presença de dehidro-epideoxiarteanuína B e de um outro composto (composto B). As condições fotoperiódicas de cultivo influenciaram na produção destes dois compostos, sendo que sobre condição de escuro contínuo, dehidro-epideoxiarteanuína B foi intensamente produzido e o composto B foi detectado em pequenas proporções, enquanto que sob fotoperíodo de 16 horas, o inverso ocorreu. A quantificação de dehidro-epideoxiarteanuína B por Cromatografia Gasosa acoplada a um Detector de Ionização de Chamas (CG/FID) revelou um aumento de aproximadamente cinco vezes na produção deste composto pelas raízes normais cultivadas sobre escuro contínuo em relação às raízes cultivadas na presença de 16 horas de luz. O terpeno dehidro-epideoxiarteanuína B não estava presente nas raízes transgênicas.

Genetic transformation of Eclipta prostrata by Ri T-DNA / 中草药

Object To establish the culture system of hairy root of Eclipta prostrata (L.) L.. Methods Hairy root of E. prostrata was obtained from infected cotyledon explants after infection with Agrobacterium rhizogenes strains A4, R1601 and ATCC15834, elite strains were screened and growth curves determined. The transformation of Ri T DNA was examined through high voltage paper electrophoresis. Results The hairy root was originally obtained from E.prostrata. The result of high voltage paper elctrophoresis confirmed the transformation of T DNA from Ri plasmid to the hairy root. Conclusion The acquisition of hairy root of E. prostrata provided further a foundation for the industrial production of active drug component.