• Plant Biotechnology Reports
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• 제2권3호
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• pp.171-177
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• 2008

This paper reports an improved protocol for isolation, culture and regeneration of Lotus corniculatus protoplasts. A range of parameters which influence the isolation of L. corniculatus protoplasts were investigated, i.e., enzyme combination, tissue type, incubation period and osmolarity level. Of three enzyme combinations tested, the highest yield of viable protoplasts was achieved with the combination of 2% Cellulase Onozuka RS, 1% Macerozyme R-10, 0.5% Driselase and 0.2% Pectolyase. The use of etiolated cotyledon tissue as a source for protoplast isolation proved vital in obtaining substantially higher protoplast yields than previously reported. Culture of the protoplasts on a nitrocellulose membrane with a Lolium perenne feeder-layer on the sequential series of PEL medium was highly successful in the formation of microcolonies with plating efficiencies 3-10 times greater than previous studies. Shoot regeneration and intact plants were achieved from 46% of protoplast-derived cell colonies.

• 생약학회지
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• 제26권4호
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• pp.419-425
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• 1995

Corydalis remota Fish. ex Max. (Papaveraceae) is a well known medicinal plant being used as analgesics or anticonvulsive in oriental medicine. As the alkaloid content is known to vary depending on the environmental factors, the technology of plant tissue culture can be adopted as source of Corydalis-alkaloids. The present study describes an establishment of tissue cultures of Corydalis which produce alkaloids consistently. Callus were induced from immature seeds of Corydalis remota by placing the seeds on MS static media containing NAA(0.25, 1.0 and 4.0 mg/l, respectively). The combined treatment of NAA(1.0 mg/l) with cytokinin(BAP 0.5 mg/l) improved the induction of callus. TLC scanning data followed by sequential extraction and purification revealed that the induced callus contains a significant amount of alkaloids. Cell suspension cultures were established by transferring the induced callus into the liquid media with the same condition of plant growth regulators as the callus culture.

• 식물조직배양학회지
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• 제26권3호
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• pp.157-161
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• 1999

본 연구는 글라디올러스 Topaz'를 재료로 액체진탕배양에 의한 캘러스의 증식효율을 향상시키기 위하여 수행되었다. 캘러스는 2,4-D 10mg/L가 첨가된 MS고체배지에 목자 외식체를 배양하여 유도하였다. 액체진탕배양에 의한 캘러스의 증식은 2,4-D 0.05 mg/L가 첨가된 MS배지를 사용하여 2$0^{\circ}C$, 16시간 일장하에서 배지를 100 mL 삼각플라스크에 20 mL 분주하고, 수평회전식 진탕배양기의 회전속도를 75 rpm으로 하였을 때 증식속도가 가장 빨랐다. 또한 동일 배지에서 캘러스의 계대배양 역시 가능하였다.

• 화훼연구
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• 제17권3호
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• pp.165-171
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• 2009

팔레높시스 화경의 모든 부위를 사용한 화경절편조직을 기내 배양하여 효율적이고 변이 발생률이 낮은 균일한 식물체를 일시에 대량생산하기 위하여 Hyponex와 생장조절물질 TDZ, BA의 적정 농도를 규명하고자 하였다. 두 공시 품종은 Hyponex, TDZ, BA 농도별로 floral stem, shoot, 다신초의 기관형성 발생에서 차이를 보였다. 팔레높시스의 화경절편조직 배양에 적절한 Hyponex의 농도는 $4g{\cdot}L^{-1}$ 가 가장 효과적이었으며, 다신초 형성에 효과적인 TDZ의 농도는 $0.3mg{\cdot}L^{-1}$이고, BA의 농도는 $5mg{\cdot}L^{-1}$ 이었다. 화경절편조직 배양 시 Hyponex $4g{\cdot}L^{-1}$을 기본배지로 하여 TDZ와 BA의 효과를 비교한 결과 TDZ가 더 효과적이었다.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2003년도 심포지엄
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• pp.50-58
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• 2003

Isolation and culture of leaf protoplasts from two chilli cultivars (Capsicum annuum var. accumnatum and Bird chilli) were developed to enhance selection process in the somatic hybridization programmes. In order to isolate the protoplasts from leaves of these two chilli cultivars different incubation periods (3, 5 and 10 hours) were tested with combinations of enzyme mixtures containing cellulase and macerozyme. Leaves were incubated on three enzyme mixtures (2% cellulase + 0.4% macerozyme, 1% cellulase + 0.2% macerozyme and 0.5% cellulase + 0.1 % macerozyme in 13% mannitol) at 251oC in the dark. Three hours of incubation using 2% cellulase and 0.4% macerozyme was the best for the protoplast isolation of both chilli cultivars tested. The yield was 5 ${\times}$ 108protoplasts/ml/ g leaf tissue in both chilli varieties. It was found that in the mixed nurse method using Nagata and Takebe (NT) medium supplemented with 1.0mg/12,4-D, NAA and BAP with 0.5M mannitol and 1.2% Sea Plaque agarose is the best medium for protoplast culture. Protoplasts of Capsicum annum var. accumnatum were alive for 14 days forming cell walls and initiating cell division.

• KSBB Journal
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• 제17권6호
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• pp.582-585
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• 2002

백합(Lilium longiflorum)으로부터 수집한 화분에 대하여 기내배양, Agrobacterium 및 vacuum infiltration과정을 이용한 형질전환 그리고 kanamycin배지에서의 선별배양을 통하여 PCR에 의하여 증폭된 1.7 kb의 human tissue plasminogen activator(tPA) cDNA의 발현을 분석하였다. 16시간 정도 배양한 신장화분관의 western blotting결과, human standard와 유사한 크기로의 발현을 확인하였다. 이로써 백합화분은 신속한 단백질발현의 분석을 위한 일회성 생산숙주로서의 가능성을 제시하고 있다.

• 한국작물학회지
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• 제41권6호
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• pp.704-709
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• 1996

모시풀의 경편과 흡지의 배양을 통하여 완전한 식물체를 대량증식하기 위하여 소독방법과 생장조절제 처리 효과에 대한 기내배양을 실시하여 다음과 같은 결과를 얻었다. 1. 모시풀 경편 배양시 소독은 초음파세체기를 이용한 2% NaClO를 20분 동안 하였을때 오염률이 3.3%로 가장 낮았으며, 식물체도 79%가 생존하였고, 건실한 묘를 생산할 수 있었다. 2. 생장조절제 처리효과에서는 NAA(0.02mg/$\ell$)+ BA(1.5mg/$\ell$) + GA3(0.1mg/$\ell$) 혼합처리가 캘러스 형성이 안되고, 식물체 형성률이 96%였으며, 건실한 묘를 생산할 수 있었다. 3. 치상부위별로는 흡지보다 경편배양이, 품종별로는 개량종인 서방종보다 재래종인 보성종이 증식효률이 높았다. 4. 순화 효율은 상토 배합과 호르몬 처리에 있어서 버미큐라이트 : 모래 ：황토 =1 : 1 : 1의 배합과 NAA 1000ppm을 30분간 담근 후 이식한 순화율이 99%로서 건실하였으며, 식물체는 대부분 정상이었다.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• 제30권5호
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• pp.438-443
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• 2004

Objectives : The proper development of the facial structures relies upon a sequence of tightly regulated signaling interactions between the ectoderm and mesoderm involving the participation of several families of signaling molecules. Among these, bone morphogenetic proteins (BMPs) have been suggested to be a key signal that regulates the development of the mandible and the initiation and morphogenesis of the teeth. The aim of this study was to examine the artificial development of the mandibular structures and to examine the role of BMPs on tooth morphogenesis and differentiation using an organ culture system. Materials and Methods : The tooth germs from Ed 11.5, 13.5 mice were dissected, and transplanted into the diastema of the mandible primordia. The mandibles containing the transplanted tooth germs were cultured in vitro. During this period, beads soaked with BMP4 were implanted around the transplanted tooth germs. In addition, a diastema block containing the transplanted tooth germ was dissected, then transferred to an adult mouse kidney. After the organ culture, the developing mandibular explant was removed from the kidney and prepared for the tissue specimens. Odontogeneis of the transplanted tooth germs was examined after Hematoxylin-eosin, Masson-trichrome staining. Results : Proliferation and differentiation of the tooth germs cultured in the diastema was observed. In the BMP4-treated tooth germs, the formation of the first and second molars was noted. The crown of the developing tooth showed the formation of a mature cusp with the deposition of enamel and dentin matrix. In conclusion, it was confirmed that BMP4 is involved in the formation of a dental crown and the differentiation of ameloblasts and odontoblasts of the molar tooth during the development of the transplanted tooth germs.

• Asian-Australasian Journal of Animal Sciences
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• 제13권2호
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• pp.161-166
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• 2000

This study was designed to determine the effects of the insulin-like growth factor (IGF-1) and estradiol $17-{\beta}$ on the in vitro proliferation of stromal vascular cell from Hanwoo omental, subcutaneous, intermuscular and intramuscular adipose tissues. Cells were cultured in M199+20% newborn calf serum and the proliferation of cells was measured by direct microscopic cell counting and change of genomic DNA amount. Cell numbers increased slightly over the first 72 hour of culture and then increased greatly, regardless of adipose tissue depots. In IGF-1 treatment, the number of omental preadipocytes maintained highest level from the beginning to the 20th day of culture. However, in estradiol-$17{\beta}$ treatment, those tended to be lower than the control from the beginning of culture and significantly lower at the 24th day. When IGF-1 was added to subcutaneous preadipocytes, the numbers of cells were higher from 11th day than those from other treatments, although there was no statistical significance. For intermuscular preadipocytes treated with IGF-1, its numbers were significantly (p<0.05) higher at 11th day, and in the other days it showed a similar tendency to those of the subcutaneous tissue. In this experiment, preadipocytes were taken from 24 month old fully matured steers and the highest proliferation rate was shown in intramuscular tissue followed by those of subcutaneous preadipocytes. Addition of $5{\mu}M$ estradiol-$17{\beta}$ to the growth medium failed to promote the replication of Hanwoo preadipocytes, as indicated by direct cell counts and total genomic DNA content. As the culture period proceeded, the amounts of DNA were increased, but the patterns of increment were not consistent with the results of cell numbers.

• Tissue Engineering and Regenerative Medicine
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• 제15권6호
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• pp.781-791
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• 2018

BACKGROUND: Glucosamine hydrochloride (GlcN HCl) has been shown to inhibit cell growth and matrix synthesis, but not with N-acetyl-glucosamine (GlcNAc) supplementation. This effect might be related to an inhibition of critical growth factors (GF), or to a different metabolization of the two glucosamine derivatives. The aim of the present study was to evaluate the synergy between GlcN HCl, GlcNAc, and GF on proliferation and cartilage matrix synthesis. METHOD: Bovine chondrocytes were cultivated in monolayers for 48 h and in three-dimensional (3D) chitosan scaffolds for 30 days in perfusion bioreactors. Serum-free (SF) medium was supplemented with either growth factors (GF) $TGF-{\beta}$ ($5ng\;mL^{-1}$) and IGF-I ($10ng\;mL^{-1}$), GlcN HCl or GlcNAc at 1mM each or both. Six groups were compared according to medium supplementation: (a) SF control; (b) SF + GlcN HCl; (c) SF + GlcNAc; (d) SF + GF; (e) SF + GF + GlcN HCl; and (f) SF + GF + GlcNAc. Cell proliferation, proteoglycan, collagen I (COL1), and collagen II (COL2) synthesis were evaluated. RESULTS: The two glucosamines showed opposite effects in monolayer culture: GlcN HCl significantly reduced proliferation and GlcNAc significantly augmented cellular metabolism. In the 30 days 3D culture, the GlcN HCl added to GF stimulated cell proliferation more than when compared to GF only, but the proteoglycan synthesis was smaller than GF. However, GlcNAc added to GF improved the cell proliferation and proteoglycan synthesis more than when compared to GF and GF/GlcN HCl. The synthesis of COL1 and COL2 was observed in all groups containing GF. CONCLUSION: GlcN HCl and GlcNAc increased cell growth and stimulated COL2 synthesis in long-time 3D culture. However, only GlcNAc added to GF improved proteoglycan synthesis.