• Journal of Plant Biotechnology
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• v.30 no.1
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• pp.65-71
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• 2003

This research aims the production of anti-tumor substances through in vitro culture of hairy roots transformed by Agrobacterium rhizogenes in Artemisia sylvatica MAX and the effect of culture conditions on optimum growth of hairy roots. We investigated the optimum medium, pH, carbon source, sucrose, light, Fe and polyamine conditions of various lines of hairy roots (NK3, NK4, YX. NK3-10) induced from Artemisia sylvatica to increase the optimum growth of hairy roots. MS medium was the best for optimum growth of hairy root clone, NK3-S10. The optimum culture period was 4 weeks for NK3-S10. The optimum sucrose concentration was 3.5%. The optimum concentration of FeSO$_4$, spermine and spermidine was 0.1 mM, 10 mM and 100 mM, respectively.

• KSBB Journal
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• v.21 no.2
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• pp.123-128
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• 2006

A transgenic hairy root clone AG-04 of Astragalus membranaceus was obtained following co-cultivation of leaf explants with Agrobacterium rhizogenes ATCC15834. This clone was examined for its growth and production of cyclolanostane-type saponins, astragalosides I, II, and III, under various culture conditions. Among the five basal media tested, Shenk and Hildebrandt(SH)(18) medium was best for roots growth and astragalosides production. The maximum root biomass was obtained at inoculum size of 500 mg FRW per flask, initial sucrose concentration of 3%, and shaking speeds of 90 rpm. The astagalosides production was promoted when the hairy root clone AG-04 was cultured at shaking speeds of 120 rpm and light irradiation of 18 h. Astragaloside contents was also stimulated with high initial sucrose concentration, and the maximum astargalosides contents of 6.21 %/g DRW was obtained at initial sucrose concentration of 6%. The addition of chitosan(100 mg/L) to the culture medium was significantly increased astragalosides production. This was 2.1 times higher than that obtained in a control culture without chitosan.

• Journal of Plant Biology
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• v.39 no.1
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• pp.63-69
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• 1996

A hairy root clone of Panax ginseng C.A. Meyer, HRB-15 was cultured iu various conditions with 3 L bubble type bioreactor to enhance both growth and ginsenoside production. The hairy roots were more rapidly grown under the dark condition than under the light condition. However, total amount of ginsenoside of hairy roots cultured under the light for 30 days increased 2 folds as compared with the dark condition and was 1.10% based on 6 ginsenosides. Especially, ginsenoside-Re was significantly increased and some ginsenosides except for ginsenoside-Re was slightly reduced. Also, the growth of hairy roots decreased about 30% as compared with the dark condition. In contrast, addition of sodium acetate led to decreased production of ginsenoside and growth of hairy roots under light condition. The influence of potassium dihydrogenphosphate concentration was examined in MS medium and a 1.25 mM concentration was found to be the most appropriate for growth and ginsenoside production under light condition. Two-step process of hairy roots culture with yeast elicitation or without ammonia in culture medium was developed to enhance growth and giusenoside synthesis. $50\;\mu\textrm{g}$ of yeast elicitor per g of fresh weight showed a synergistic effect on the ginsenoside synthesis of hairy roots on 20 days after culture. At that time, the content of total ginsenoside was 1.15%, while the growth of hairy roots decreased 21 % as compared with the dark condition. In addition, when elimination of ammonia on 20 days after culture, the content of total ginsenoside was 1.26% with significant increment of ginsenoside-Rd (0.27%) in addition to ginsenoside-Re and the growth of hairy roots decreased 10% as compared with the dark condition. In this system, we have demonstrated a unique two-step process of hairy root cultures to maximize biomass and secondary metabolites. It has found possibility to enhance ginsenosides production by growing hairy roots in this method.

• Korean Journal of Medicinal Crop Science
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• v.10 no.4
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• pp.243-248
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• 2002

The effects of basal media, carbon, nitrogen, phosphate and some major macro elements on growth and shikonin production in Lithospermum erythrorhizon hairy root culture were studied. Among examined media, growth of hairy root cultured in B5 liquid medium was rapid, whereas shikonin production was high in MS liquid medium. Under B5 basal medium, sucrose concentration for optimal growth and shikonin production was 9% and 4% respectively. The growth and shikonin production on pH changes in B5 medium resulted little effect in pH 5.8 to pH 8.8 ranges, whereas growth was decreased dramatically in both above 8.8 and under 5.8. Nitrogen source and concentration effected on the growth and shikonin production. The highest growth rate was in B5 medium (50 mM $KNO_3$ and 1 mM $NaH_2PO_4)$, whereas the highest shikonin production was in the condition supplemented with 5 mM $KNO_3$ and 10 mM $NaH_2PO_4$.

• Korean Journal of Plant Tissue Culture
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• v.27 no.6
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• pp.485-489
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• 2000

The patterns and contents of ginsenosides were examined in normal root parts and hairy root lines of Panax ginseng C. A. Meyer. Ginsenoside-Rb$_1$, -Rb$_2$, -Rc, -Rd, -Re, -Rf, -Rg$_1$, -Rg$_2$ were detected in normal roots and hairy roots of ginseng. The patterns and contents of ginsenosides in that were very difference each other. The contents of total ginsenoside of hairy root (KGHR-1) was 17.42 mg/g dry wt, it's highest compared to others. Ginsenoside contents of hairy root (KGHR-1) was higher on ginsenoside-Rd, Rg$_1$, KGHR-5 was higher on ginsenoside-Rb$_1$, Rg$_1$, and KGHR-8 was higher on ginsenoside-Rd, Re than others. The contents of total ginsenosides on 6 years old ginseng cultured in the field were high in the order of main root, lateral root and fine roots, and content of ginsenosides in fine roots was 3.2 times higher than that in main root. The ratio of ginsenoside-Rg$_1$to total ginsenosides were about 3.43%, 8.68% and 14.18% respectively on fine root, lateral root and main root, it's very lower than that in hairy roots. It is suggested that specific ginsenosides can be produce in cultures of ginseng hairy roots.

• 한국생물공학회:학술대회논문집
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• 2005.10a
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• pp.417-420
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• 2005

Plants generally produce secondary metabolites in nature as a defense mechanism against pathogenic and insect attack. In this study, we applied several abiotic elicitors in order to enhance growth and ginseng saponin biosynthesis in the hairy roots of P. ginseng. Generally, elicitor treatments were found to inhibit the growth of the hairy roots, although simultaneously enhancing ginseng saponin biosynthesis. The addition of selenium at inoculum time did not significantly affect ginseng saponin biosynthesis. However, when 0.5 mM selenium was added as an elicitor after 21 days of culture, ginseng saponin content and productivity increased to about 1.31 and 1.33 times control levels, respectively. These results suggest that processing time for the generation of ginseng saponin in a hairy root culture can be reduced via the application of an elicitor.

• Journal of Plant Biotechnology
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• v.34 no.4
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• pp.285-291
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• 2007

This study describes conditions for the mass production of activation-tagged mutant hairy root lines of ginseng by cocultivation with Agrobacterium rhizogenes. Because it is not currently possible to produce progeny from transgenic ginseng, a loss-of-function approach for functional genomics cannot be appliable to this species. A gain-of-function approach is alternatively the choice and hairy root production by cocultivation of A. rhizogenes would be most practical to obtain a large number of mutants. Various sources of explants were subjected to genetic transformation with various strains of A. rhizogenes harboring the activation-tagging vector pKH01 to determine optimum conditions for the highest frequency of hairy root formation on explants. Petiole explants cocultivated with A. rhizogenes R1000 produced hairy roots at a frequency of 85.9% after 4 weeks of culture. Conditions for maximum growth or branching rate of hairy roots were also investigated by using various culture media. Petiole explants cultured on half strength Schenk and Hildebrandt medium produced vigorously growing branched roots at a rate of 2.6 after 4 weeks of culture. A total of 1,989 lines of hairy root mutants were established in this study. These hairy root lines will be useful to determine functions of genes for biosynthesis of ginsenosides.

• Korean Journal of Plant Tissue Culture
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• v.28 no.6
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• pp.347-351
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• 2001

Panax ginseng C.A. Meyer is a well-known Korean traditional medicine. Until now, even though major research of ginseng has been focused on the pharmacological effect, clinical application and chemical analysis of extracted secondary metabolite for several years, the physiology and gene functions of ginseng were not well known. In this research, we have developed the protein extraction methods of ginseng root and hairy root for proteome analysis in order to elucidate the gene(s) function of ginseng. Using the liquid nitrogen (equation omitted) TCA method as protein extraction method, about 660 protein spots were detected on the 2-DE gel of hairy root. Additionally, comparative analysis result of 2-DEs of ginseng root (equation omitted) hairy root suggested that proteomes of same organism could be changeable according to the culture condition, growth stages and other stimulus.

• KSBB Journal
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• v.10 no.4
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• pp.435-440
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• 1995

During culture the effect of polyamine on hairy root of Buplerum falcatum by infection of Agrobacterium rhizogenes was studied. The fresh and dry weight of hairy root which cultured for 3months in MS medium increased about 7-fold when spermine at 10${\mu}$M, 100${\mu}$M was treated. After suspension culture of B. falcatum in MS medium containing putrescine(10${\mu}$M, 100${\mu}$M) or spermine(10${\mu}$M), the contents of endogenous polyamine (putresclne, spermidine, spermine) was higher than that of control. The ${\beta}$-glucan synthetase II activity by polyamine treatment was increased： especially spermidine (100${\mu}$M) and spermine(100${\mu}$M) stimulated it by about 180% and 220% respectively. These results suggest a possible role of polyamine as growth regulator in B. falcatum hairy root cultures.

• Journal of Ginseng Research
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• v.27 no.2
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• pp.78-85
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• 2003

By the Agrobacterium rhizogenes A$_4$ were induced a transformed callus of ginseng hairy root and examine to find the possibility whether it can produce certain ginsenoside. Investigations for a finding out to optimal culture medium showed that BA application is better than more factorial composition between auxins and cytokinins. For the induction of hairy root callus of ginseng, l/2 MS medium containing 1 to 3 mg of benzyladenine(BA) per liter gave the best result. The growth of ginseng hairy root callus(GHC) cultured with the 1/2MS medium supplemented with 2 mg BA/L was selected for best suspension cultures. The optimum concentration of BA for ginsenosides production was found to be 2 mg/L. Probably the inoculum size of callus plays a role with the ginsenoside production in suspension culture. AS for inoculum size of callus, 50 mg was superior to 150 mg for growth and ginsenoside production. Ginsenoside contents were highest in the suspension culture grown for four weeks under continuous light condition. In fact that continous light treatment promote strongly the synthesis of ginsenoside of the hairy root callus is first result in the world and the numerously induced root hairs of the callus leads a new method for ginsenoside production.