• Journal of Plant Biotechnology
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• 제32권4호
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• pp.313-319
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• 2005

다양한 탄소원을 사용하였을 시, 세포성장에 미치는 영향은 배양 5일제 glucose를 사용한 배양세포에서 dry cell weight가 11.2 g/L, wet cell weight가 310.8 g/L로 가장 높았다. 이후 배양 10일차에는 오히려 sucrose를 탄소원으로 사용한 배양세포에서 dry cell weight가 13.4 g/L, wet cell weight가 480 g/L로 가장 높이 성장한 것을 확인할 수 있었다. Total secreted protein의 경우는 배양 10일차에 sucrose를 기본 탄소원으로 사용하였을 때 110.3 mg/L로 가장 높게 나왔다. Total secreted protease 역시, 배양 10일차에 sucrose를 기본 탄소원으로 사용한 배양씩에서 3950 U/L로 가장 많은 양이 측정되었다. 최종적으로 재조합된 단백질인 hGM-CSF의 생산량에 대한 측정결과, sucrose를 기본 탄소원로 사용한 배지에서 배양 10일차에 56 mg/L로 가장 높게 측정됨을 확인할 수 있었다. 이 외에 다양한 탄소원 농도변화에 따른 total secreted protein과 hGM-CSF의 안정성 평가를 확인해본 결과, total secreted protein의 경우, 비교대상으로 사용한 sucrose에서는 농도변화에도 큰 손실률을 보이지는 않았으나, sorbitol, mannitol, fructose, glucose의 경우에는 농도가 높아질수록 배양액내의 protein의 손실률이 증가 하였다. 총 분비된 hGM-CSF 역시 sucrose에 비해 sorbitol, mannitol, fructose, glucose에서는 분비된 hGM-CSF의 양이 크게 감소하는 것을 확인할 수 있었고, 탄소원의 농도가 높아질수록 hGM-CSF의 양이 감소하는 경향이 크게 나타났다.

• Plant Biotechnology Reports
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• 제5권4호
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• pp.367-373
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• 2011

Linum album accumulates anti-tumor podophyllotoxin (PTOX) and its related lignans, which were originally isolated from an endangered species Podophyllum. In the present study, we examined the effects of five fungal extracts on the production of lignans in L. album cell cultures. Fusarium graminearum extract induced the highest increase of PTOX [$143{\mu}g\;g^{-1}$ dry weight (DW) of the L. album cell culture], while Rhizopus stolonifer extract enhanced the accumulation of lariciresinol up to $364{\mu}g\;g^{-1}$ DW, instead of PTOX. Typical elicitors, such as chitin, chitosan, or methyl jasmonate (MeJA), were shown to be less effective in lignan production in L. album cell cultures. These results verified the advantages of fungal extracts to increase lignan production in L. album cell culture, and suggested potential on-demand metabolic engineering of lignan biosynthesis using differential fungal extracts.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2001년도 추계학술발표대회
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• pp.51-51
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• 2001

• Applied Biological Chemistry
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• 제48권4호
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• pp.425-430
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• 2005

[${\beta}-sitosterol$]은 식물 스테롤로서 인간의 전립선암과 대장암 세포의 성장을 억제하고 생체내 콜레스테롤 농도를 감소시킨다. 본 연구에서는 쑥갓세포 배양에서 ${\beta}-sitosterol$ 생산의 최적화 연구를 수행하였다. 그래서 쑥갓(Chrysanthemum coronarium L.)으로부터 캘러스 유도는 NAA와 BAP의 농도가 각각 1 mg/l의 조합에서 최적이었으며 이들 캘러스로부터 현탁배양 세포주를 확립하였다. 현탁 배양시 초기 세포농도 2 g DCW/l에서 조성이 각각 1배인 탄소원(30 mg/l), 질소원(1900 mg/l $KNO_3$, 1650mg/l $NH_4NO_3$), 무기인산원(170 mg/l)을 포함하는 MS 배지에서 ${\beta}-sitosterol$ 생산이 최적으로 나타났다. Shake-flask를 이용한 현탁배양에서 ${\beta}-sitosterol$의 최대 생산량은 $150{\mu}g/g$ DCW이었다. 그리고 공기부유식 생물반응기의 배양에서는 100 cc/ml의 통기량에서 ${\beta}-sitosterol$의 생산이 $142.8{\mu}g/g$ DCW으로 나타났다.

• Plant Biotechnology Reports
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• 제5권3호
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• pp.245-254
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• 2011

This study describes an efficient protocol for Agrobacterium tumefaciens-mediated transformation of two subgroups of genotype AAA bananas (Musa acuminata cv. Pei Chiao and Musa acuminata cv. Gros Michel). Instead of using suspension cells, cauliflower-like bud clumps, also known as multiple bud clumps (MBC), were induced from sucker buds on MS medium containing $N^6$-Benzylaminopurine (BA), Thidiazuron (TDZ), and Paclobutrazol (PP333). Bud slices were co-cultivated with A. tumefaciens C58C1 or EHA105 that carry a plasmid containing Arabidopsis root-type ferredoxin gene (Atfd3) and a plant ferredoxin-like protein (pflp) gene, respectively. These two strains showed differences in transformation efficiency. The EHA105 strain was more sensitive in Pei Chiao, 51.3% bud slices were pflp-transformed, and 12.6% slices were Atfd3-transformed. Gros Michel was susceptible to C58C1 and the transformation efficiency is 4.4% for pflp and 13.1% for Atfd3. Additionally, gene integration of the putative pflp was confirmed by Southern blot. Resulting from the pathogen inoculation assay, we found that the pflp transgenic banana exhibited resistance to Fusarium oxysporum f. sp. cubense tropical race 4. This protocol is highly advantageous to banana cultivars that have difficulties in setting up suspension cultures for the purpose of quality improvement through genetic transformation. In addition, this protocol would save at least 6 months in obtaining explants for transformation and reduce labor for weekly subculture in embryogenic cell suspension culture systems.

• Journal of Plant Biotechnology
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• 제3권2호
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• pp.83-88
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• 2001

A cell culture system of poplar (Populus alba x P.glandulosa) was established to test four different methods for evaluation of cellular stresses. Two different kinds of stresses were given to the cultures by adding either Pb(NO$_3$)$_2$ or paraquat and the cellular responses were monitored during a week period. While fresh weight reduction was observable in two days after the treatment of Pb(NO$_3$)$_2$, such changes were apparent only in later stage in paraquat treated cultures. Cells in paraquat treated cultures in the first 3 days showed no alteration in fresh weight as compared to untreated cultures, but had their MTT reducing activities completely inhibited. Neither Evans blue staining nor ion conductivity of the medium was consistent with fresh weight changes of the cultures. Overall, cell clumps formed during suspension culture appeared to interfere with staining and washing reactions and thus cause the assays unreliable. Among the four methods examined, fresh weight changes and MTT reducing activity appeared to be the most reliable and consistent.

• 한국약용작물학회지
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• 제15권5호
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• pp.346-351
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• 2007

An efficient somatic embryogenesis and plant regeneration protocol was developed for Schisandra chinensis Baill, using embryogenic cell suspensions and optimized media conditions. Friable embryogenic callus was induced from cotyledonary leaf and hypocotyl explants of 7 days old seedlings on MS agar medium supplemented with 1.0 to $4.0\;mg\;l^{-1}$ of 2,4-dichlorophenoxyacetic acid (2,4-D). Fast growing and well dispersed embryogenic cell suspensions were developed within two months when embryogenic calli were transferred to MS liquid medium containing $1.0\;mg\;l^{-1}\;2,4-D$. One third strength of MS medium was the best for both overall growth and development of somatic embryos in liquid culture. Over 3400 viable somatic embryos were produced from each 150 ml flask with an initial cell density of 30 mg in 30 ml medium. Germinated somatic embryos developed in liquid medium converted into plantlets after transferred to half-strength MS semi-solid medium. Approximately 90% of the converted plantlets were successfully transplanted to soil and grew into fertile plants.

• Journal of Plant Biotechnology
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• 제31권2호
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• pp.163-167
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• 2004

본 연구에서는 hGM-CSF 유전자가 도입된 형질전환 담배세포에서 sucrose의 농도가 배양 배지내 분비된 hGM-CSF, 총분비 단백질, 그리고 단백질 분해효소에 미치는 효과를 보았다. 10, 30, 60, 90, 120, 150 g/L의 sucrose 농도로 배양한 결과 sucrose농도가 30g/L일 때 건조중량은 11.22g/L, 분비된 hGM-CSF의 양은 181.53 $\mu\textrm{g}$/L, 그리고 총단백질은 66.8 mg/L.로 시료채취 5일 때 가장 높은 값을 나타내었고, 단백질 분해효소는 sucrose농도가 120 g/L일때 2660 U/L.의 값을 나타내었다. 그러나 배양 10일째 sucrose농도 60 g/L에서 건조중량이 28.36g/L로 가장 높은 값을 나타낸 반면 분비된 hGM-CSF의 최대 값은 95 $\mu\textrm{g}$/L.로 sucrose농도가 150 g/L일 때 가장 높은 값을 나타내었다.

• Journal of Plant Biology
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• 제31권2호
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• pp.91-100
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• 1988

The effect of Ca2+ and polyamines on the activity $\beta$-glucan synthetase II(GSII) related to cell wall synthesis was studied in carrot suspension cultured cells. The activity of GS II is four times higher than that of $\beta$-glucan synthetase I in carrot suspension cultured cells and in vitro expreiment, the activity of GSII was increased in response to increase in concentration of Ca2+ and polyamines. When carrot suspension cultured cells were incubated together with Ca2+ and polyamines, the GSII activity was high at 0.1mM of Ca2+ and 1mM of putrescine. Also, polycationic poly-L-lysine and poly-L-ornithine increased about 50% the GSII activity than that of the control, respectively. These results may imply that Ca2+ and polyamines were related to the enzyme activity as a polycationic nature. In addition, verapamil as the calcium channel blocker and flunarizine as an antagonist of calcium mechanism in cytoplasm decreased GSII activity ramarkably, Ca2+ and calmodulin stimulated GSII activity as Ca2+ of free ion rather than Ca2+ calmodulin complex. The effect of 2,4-D on the GSII activity in culture medium is shown to be low at 0.1mg per liter and GSII activity increased about 30% more than that of the 0.1mg/l at the range of 0.3-1.0mg per litere. Cummulative results suggest that Ca2+ and polyfamines stimulate the cell wall synthesis by means of the enhancement of GSII activity responsible for synthesizing the cell wall components.

• 식물조직배양학회지
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• 제21권3호
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• pp.183-186
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• 1994

태백벼의 현탁배양세포주로부터 유도된 체세포배를 낱개로 알진산 캡슐화하여 인공종자화하였다. 인공종자는 1/2 MS 고체배지에서 73%의 발아율을 나타내었으며 알진산 캡슐은 체세포배의 발아율에 영향을 주지 않았다. 그러나 멸균되지 않은 여과지에서는 발아율이 60%로 낮아졌다. 캡슐화된 진정종자는 무균상태여부에 관계없이 높은 발아율을 나타내었다. 이상의 결과로 무균상태가 유지되지 않을 때 인공종자 발아율이 낮아지는 것은 체세포배가 진정종자의 접합자배보다 오염을 견디기 어렵기 때문인 것으로 사료된다.