• KOREAN JOURNAL OF CROP SCIENCE
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• v.42 no.5
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• pp.615-625
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• 1997

An efficient protocol for plant regeneration from protoplasts of the indica rice variety IR43 has been developed. The procedure involved plating of embryogenic suspension-derived protoplasts on the surface of a filter membrane overlaying agarose-embedded feeder cells. Lolium multiflorum cell suspensions were preferable to these of Oryza ridleyi as feeder cells and Lolium suspensions supported colony formation from up to 0.68％ of the protoplasts, depending on the age of cell suspensions. Plant regeneration frequency was significantly improved by using maltose alone or in a 1:1(w/w) combination with sucrose as carbohydrate source and a simple dehydration treatment using a high concentration of agarose in the regeneration medium. Medium containing maltose or maltose mixed with sucrose increased the plant regeneration frequency compared with medium containing sucrose alone. The plant regeneration frequency was increased to 30.7 to 70.7％ following dehydration treatment, while the non-treated controls showed a regeneration frequency of 3.1 to 30.6％. Protoplast-derived plants were transferred to the glasshouse, flowered with morphologically normal.

• Korean Journal of Plant Tissue Culture
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• v.21 no.2
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• pp.117-121
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• 1994

The optimal level of growth regulator for callus initiation stem explants was BAP 0.1mg/L combined with 2,4-D 1.0 mg/L in Murashige and Skoog (MS) medium supplemented with 30g/L sucrose and 10g/L agar, and that for cell growth was BAP 0.1+2,4-D 0.5 mg/L in MS liquid medium. The regeneration frequency of P. oleracea cells was significantly increased by subjecting the cells to dessication for 1and 2 h up to 83%, respectively, as compared with untreated control showing 61%. Cell viability and survival rate was inhibited by pretreatment of 0.6% NaCl for 2 days, while regeneration ability was not affected by the treatment. Pretreatment of 100g/L sucrose for 2 days markedly stimulated the regeneration of cells up to 81%. These results suggest that in addition to physiological changes, water stress resulted from dessication and high concentration of sucrose and NaCl is closely related to the regeneration of P. oleracea cultured cells.

• Journal of Plant Biotechnology
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• v.4 no.3
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• pp.129-134
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• 2002

We report the development of a new selection system for the transformation of pepper plants by mannose. In order to achieve this, we first tested several factors related to regeneration conditions. Among the 30 inbred lines examined, line P9l5 was able to generate shoots at the highest rate from both cotyledons and hyporotyls in MS media. A dosage curve for optimizing the selection conditions was established by mixing mannose (range 0-50 g/L) and sucrose (range 0-30 g/L). The least selection pressure on shoot formation was created by a mixture of sucrose and mannose at 20 g/L and 15 g/L, respectively, and the threshold for ultimate tissue death was 50 g/L of mannose irrespective of the sucrose concentration. However, we found that mannose itself was not the sole inhibitor of pepper shoot development. High concentrations of sucrose (30 g/L) contributed additively to the inhibition of shoot formation at higher mannose concentrations. Genotype preference is a major factor that enhances regeneration ability in mannose media, as was observed in MS media. P9l5 and P410 line had high regeneration rates under mannose selection conditions in the presence of Agrobacterium infection. Different virulence levels of Agrobacterium strains did change the regeneration rates, probably due to interaction with the specificities of the inbred lines. Taken together, P9l5 offers the best pepper inbred line for transformation and we recommend a selection condition of 20 g/L of sucrose and 15 g/L or more of mannose up to 50 g/L in media.

• Korean Journal of Microbiology
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• v.24 no.4
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• pp.364-369
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• 1986

A procedure for the preparation, regeneration and fusion of protoplasts of Streptomyces tubercidicus was confirmed. Also, protoplast releasingprocesses from mycelia were observed by scanning electron microscope. Three types of protoplasts releasing processes-from the hyphal tip, hyphal end regions and lateral regions of the hyphae-were observed. More than 17% regeneration efficiency was obtained by regeneration medium that is composed of tryptone-yeast extract-sodium acetate-$MgCl_2-CaCl_2$-sucrose. Optimal concentrations of $Ca^{++},\;Mg^{++}$ and sucrose in the regeneration medium were 50mM, 0.4-0.5M respectively. Above 30% of fusion frequency of the protoplasts derived from two auxotrophic strains of S. tubercidicus was induced by polyethylene glycol 4000(60% w/v).

• Microbiology and Biotechnology Letters
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• v.14 no.3
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• pp.251-257
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• 1986

Optimum conditions for the protoplast formation and regeneration of Lactobacillus casei have been searched for. L. casei cells were converted to protoplast by treating with 10$\mu\textrm{g}$/$m{\ell}$ of mutanolysin in 20mM potassium phosphate buffer (pH6.8) containing 6mM CaCl$_2$, 6mM MgCl$_2$ and 1M sucrose. Maximum number of protoplasts was obtained when cells were taken from Tomochika's medium containing 0.5% glycine at the middle to late logarithmic growth phase. Regeneration was efficiently accomplished on the regeneration medium containing 6mM CaCl$_2$, 6mM MgCl$_2$, 0.8M sucrose and 10% of horse serum. The efficiently of the cell wall regeneration from protoplasts was 2-5% after 3-4 days of incubation at 3$0^{\circ}C$.

• Journal of Plant Biotechnology
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• v.41 no.4
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• pp.223-228
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• 2014

To establish an efficient proliferation and regeneration of PLBs (protocorm-like bodies) of Phalaenopsis plants, a variety of propagation medium types, various concentraions of sucrose as well as liquid and solid type were tested in this study. Further, activated charcoal, citric acid and ascorbic acid were compared whether these agents are suppose to reduce the browning in culture process using PLBs of Phalanopsis plants. With regard to the proper propagation medium, VW medium showed 1.3 ~ 2 times highr than those of other medium in an index of increasing for fresh weight and 50% higher than those of other medium in the frequency of shoot regeneration. However, regarding liquid and solid types of culture, there were no significant differences in the proliferation of PLBs and regeneration of shoots from PLBs. In the experiment for a variety of sucrose concentrations (0 ~ 50 g/l), 10 g of sucrose showed 30 ~ 50% higher than other concentrations in increasing index and 10 ~ 50% higher in the regeneration of shoots from PLBs. Regarding the reduction of browning in tissue culture via PLBs of Phalaenopsis plants, 1 g of activated charcoal showed only 1.5% browning of PLBs cultured. Whereas, other treatments including citric acid and ascorbic acid showed 6 ~ 16% of browning of PLBs. Therefore, activated charcoal was selected as an efficient anti-browning agents for the culture of PLBs in Phalaenopsis plants. Using above-described results can be contibuted to the establishment of mass propagation system using PLBs of Phalaenopsis plants in the future.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.49 no.1
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• pp.52-60
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• 2004

The optimized in vitro culture system was investigated for improvement of regeneration efficiencies by observing the responses of scutella-derived callus of Korean rice (Oryza sativa L.). Large variations of callus induction (43.9-93.9%) and shoot regeneration (0-88.7%) were observed among the rice cultivars depending on medium. However, shoot regeneration was significantly improved by selected utilization of basal medium, growth regulators, and carbon sources. N6 basal medium was more efficient for embryogenic callus induction than MS or LS basal medium, while MS was superior to N6 for shoot regeneration. The calli of highly regenerative cultivars grew faster and showed higher rates of green tissue formation (GT) and shoot regeneration (SR) and lower rate of callus browning (CB) than those of recalcitrant cultivars. Although a higher level of kinetin stimulated the GT and SR in highly regenerative cultivars, $10\textrm{mgL}^{-1}$ kinetin generally suppressed the GT and SR, while CB was accelerated compared to $2\textrm{mgL}^{-1}$ kinetin. Additional benefits of sorbitol combined with maltose (or sucrose) under $5\textrm{mgL}^{-1}$ kinetin were certainly confirmed on regeneration efficiencies compared to sucrose alone as carbon source and osmotic regulator. This combination showed high rate of GT and SR with multiple shoots while low rate of CB. With MSRK5SM-Pr medium ($5\textrm{mgL}^{-1}$ kinetin, 3% sorbitol, 2% maltose, $500\textrm{mgL}^{-1}$ proline), the regeneration efficiencies of total 17 out of 24 cultivars were practically improved 160% on average compared to MSRK2S ($2\textrm{mgL}^{-1}$ kinetin, 3% sucrose) control medium. Especially, the medium was most effective to the cultivars showing a medium level of regenerability such as Daesanbyeo and Dongjinbyeo and Suwon477, enhancing efficiencies more than 300-600% compared to MSRK2S medium.

• Journal of Plant Biotechnology
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• v.45 no.2
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• pp.131-139
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• 2018

We examined the effect of carbon sources on the regeneration and ex vitro acclimatization of Lycopus lucidus Turcz. ex Benth. Plantlets were regenerated on the 1/2MS medium supplemented with different concentrations (3 ~ 10%) of sucrose and glucose. The sucrose concentrations of 3% and 5% that were supplied enhanced shoot multiplication and rooting but hampered high concentration growth (including the length of the shoot and root). During ex vitro acclimatization, the tuberization of the root, the root length, the shoot length and the survival rate of Lycopus lucidus plantlets grown using 3% and 5% sucrose were found to be better than the other carbon sources and concentrations. Thus a sucrose concentration of 3% and 5% in the 1/2MS medium appeared to be better for both in vitro growth and ex vitro acclimatization of Lycopus lucidus.

• Journal of Plant Biotechnology
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• v.30 no.1
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• pp.41-45
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• 2003

A study was under taken to investigate the appropriate explant sources of flower organ and suitable cultural conditions such as light, temperature, and sucrose in plant regeneration of Iris ensata culture. Explants of perianth, ovary, pedicel, and peduncle of Iris ensata were cultured at different daylength (0, 8, 16, 24 hour), different temperatures (10, 15, 25, 3$0^{\circ}C$), and sucrose concentrations (1, 3, 6, 9%) on MS medium. Formation of adventitious roots from explants of Iris ensata was effective in the dark, while that of adventitous shoots was effective in the light. The optimum daylength for young plant regeneration was 16 hours. The optimum temperature for shoot formation of Iris ensata explants was $25^{\circ}C$ but the formation at 10 and 15$^{\circ}C$ was ineffective. Especi-ally, perianth and ovary was effective in shoot formation from flower organ expants. T-he optimum concentration of sucrose for shoots and roots formation of Iris ensata explants was 3 and 6%, respectively.

• Journal of Microbiology and Biotechnology
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• v.1 no.4
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• pp.246-250
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• 1991

Conditions for the production and regeneration in Lactobacillus bulgaricus protoplasts were investigated. Protoplasts of L bulgaricus strains were obtained by treatment with mutanolysin and lysozyme together in a protoplast forming buffer containing 0.02 M N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES) (pH 7.0) and 0.5 M sucrose. High protoplast yield was obtained from cells cultured in the de Man, Rogosa and Sharpe(MRS) medium at the middle to late logarithmic growth phase. Regeneration was efficiently accomplished with a complex medium containing 1% sucrose, 20 mM $MgCl_2$, 5% gelatin, and 0.5% bovine serum albumin. The frequency of regeneration of protoplasts was 10~20% after 5 days of incubation at $30^{\circ}C$.