• Title/Summary/Keyword: In vitro regeneration

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In Vitro and In Vivo Evaluation of Composite Scaffold of BCP, Bioglass and Gelatin for Bone Tissue Engineering

  • Kim, Woo Seok;Nath, Subrata Deb;Bae, Jun Sang;Padalhin, Andrew;Kim, Boram;Song, Myeong Jin;Min, Young Ki
    • Korean Journal of Materials Research
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    • v.24 no.6
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    • pp.310-318
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    • 2014
  • In this experiment, a highly porous scaffold of biphasic calcium phosphate (BCP) was prepared using the spongereplica method. The BCP scaffold was coated with 58S bioactive glass (BG) and sintered for a second time. The resulting scaffold was coated with gelatin (Gel) and cross-linked with [3-(3-dimethyl aminopropyl) carbodiimide] and N-Hydroxysuccinamide (EDC-NHS). The initial average pore size of the scaffold ranged from 300 to $700{\mu}m$, with more than 85 % porosity. The coating of BG and Gel had a significant effect on the scaffold-pore size, decreasing scaffold porosity while increasing mechanical strength. The material and surface properties were evaluated by means of several experiments involving scanning electron microscopy (SEM), energy-dispersive X-ray (EDX) and X-ray diffraction (XRD). Cytotoxicity was evaluated using MTT assay and confocal imaging of MC3T3-E1 pre-osteoblast cells cultured in vitro. Three types of scaffold (BCP, BCP-BG and BCP-BG-Gel) were implanted in a rat skull for in vivo evaluation. After 8 weeks of implantation, bone regeneration occurred in all three types of sample. Interestingly, regeneration was found to be greater (geometrically and physiologically) for neat BCP scaffolds than for two other kinds of composite scaffolds. However, the other two types of scaffolds were still better than the control (i.e., defect without treatment).

The effect of bacterial cellulose membrane compared with collagen membrane on guided bone regeneration

  • Lee, So-Hyoun;Lim, Youn-Mook;Jeong, Sung In;An, Sung-Jun;Kang, Seong-Soo;Jeong, Chang-Mo;Huh, Jung-Bo
    • The Journal of Advanced Prosthodontics
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    • v.7 no.6
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    • pp.484-495
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    • 2015
  • PURPOSE. This study was to evaluate the effects of bacterial cellulose (BC) membranes as a barrier membrane on guided bone regeneration (GBR) in comparison with those of the resorbable collagen membranes. MATERIALS AND METHODS. BC membranes were fabricated using biomimetic technology. Surface properties were analyzed, Mechanical properties were measured, in vitro cell proliferation test were performed with NIH3T3 cells and in vivo study were performed with rat calvarial defect and histomorphometric analysis was done. The Mann-Whitney U test and the Wilcoxon signed rank test was used (${\alpha}<.05$). RESULTS. BC membrane showed significantly higher mechanical properties such as wet tensile strength than collagen membrane and represented a three-dimensional multilayered structure cross-linked by nano-fibers with 60 % porosity. In vitro study, cell adhesion and proliferation were observed on BC membrane. However, morphology of the cells was found to be less differentiated, and the cell proliferation rate was lower than those of the cells on collagen membrane. In vivo study, the grafted BC membrane did not induce inflammatory response, and maintained adequate space for bone regeneration. An amount of new bone formation in defect region loaded with BC membrane was significantly similar to that of collagen membrane application. CONCLUSION. BC membrane has potential to be used as a barrier membrane, and efficacy of the membrane on GBR is comparable to that of collagen membrane.

In vitro Regeneration of Phragmites australis through Embryogenic Cultures

  • Lee Jeong-Sun;Kim Chang-Kyun;Kim In-Sung;Lee Eun-Ju;Choi Hong-Keun
    • Journal of Plant Biotechnology
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    • v.8 no.1
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    • pp.21-25
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    • 2006
  • Phragmites australis (reed) has received much attention as being one of the principle emergent aquatic plants for treating industrial and civil wastewater. Plant regeneration via plant tissue culture in p. australis was investigated. Three types of callus were identified from seeds on N6 medium plus 4.5 UM 2,4-dichlorophenoxyacetic acid (2,4-D). Yellow compact type showed the best redifferentiation, whereas white compact type and yellow friable were not competent to differentiate into plane. Solid medium culture was better than liquid suspension culture for enhancing callus growth when N6 medium supplemented with 4.5 ${\mu}M$ 2,4-D was used. Phytagel, as a gelling agent, was superior to agar in plant regeneration on N6 medium, supplemented with 9.4 ${\mu}M$ kinetin and 0.54 ${\mu}M$ $\alpha$-naphthaleneacetic acid (NAA). Transfer of the plantlets regenerated from kinetin and NAA-supplemented N6 medium to growth regulator-free MS medium enhanced the further development of the plantlets. Plantlets on subsequently grown to maturity when tansferred to potting soil. The regenerated plants exhibited morphologically normal. The system for plant regeneration of P. australis enables to propagate elite lines on a large scale for water purification in the ecosystem

A study of the effects of electric current on the mineralzation of the cultured calvaria bone cells (전기적 자극이 배양 두개관 골세포의 석회화에 미치는 영향에 관한 연구)

  • Park, Joon-Bong;Hur, In-Sik;Lee, Hye-Ja;Choi, Young-Chul
    • Journal of Periodontal and Implant Science
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    • v.27 no.4
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    • pp.949-961
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    • 1997
  • To date, various clinical procedures have been used to restore periodontal apparatus destroyed by periodontal disease. And then, many experimental approaches have been proceeded to develop treatment methods to promote periodontal regeneration. Mechanical, chemical treatments to enhance the attachment of periodontal tissue cells as changing the physical properties of root surfaces, bone graft procedure, and treatments for guided tissue regeneration have been used for periodontal regeneration. However, recent studies have revealed that biologic factors such as growth factors promote biologic mechanism associated with periodontal regeneration. This study was done to enucleate how ELF stimulus affect the periodontal regeneration. We can have following conclusions from this experimental results. The influence of low frequency(ELF) electric stimulus (30Hz at $lO{\mu}A$) known to promote bone formation in vivo, was evaluated for its ability to affect bone cell function in vitro. After 12 hour exposure of ELF stimulus at most appropriate densities ($5{\times}10^4\;cells/cm^2$) to increase osteoblastic cells normally, rat calvarial cells were incubated for 60 hours were used in this study. We have found ELF stimulus suppress calvarial cell proliferation and the ability of protein synthesis, enhance the alkaline phosphatase activity significantly.

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In vitro plant regeneration from axillary buds of Hibiscus syriacus L.

  • Jeon, Seo-Bum;Kang, Seung-Won;Kim, Wan-Soon;Lee, Gung-Pyo;Kim, Sun-Hyung;Seo, Sang-Gyu
    • Journal of Plant Biotechnology
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    • v.36 no.2
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    • pp.174-178
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    • 2009
  • Presently, we report a simple, reproducible and high frequency plant regeneration in Hibiscus syriacus L. using axillary buds. H. syriacus was regenerated from axillary buds directly or through a callus phase. Regenerated shoots were directly induced from young and fresh axillary buds cultured on Murashige and Skoog medium (MS) supplemented with 0.01 mg/L of the growth regulator thidiazuron (TDZ) after 2 weeks of culture. Directly induced shoots were transferred to hormone-free MS medium and root development was observed after 6 weeks. On the other hand, old and stale axillary buds were regenerated to shoots via callus induction on MS medium containing 0.01–2 mg/L TDZ after 4 weeks. A TDZ concentration of 0.01 mg/L was most effective in callus formation. Green callus was transferred to MS medium containing 0.01 mg/L α-naphthalene acetic acid (NAA) and 0.5 mg/L benzylaminopurine (BA). After 4 weeks, callus had developed into multiple shoots. Plantlets were formed from 10 week cultures of single shoots on hormone-free MS medium. Regenerated plantlets were cultured on MS medium for one month and then transferred to pots containing garden soil. Potted plants were acclimatized for one month and grown to maturity under greenhouse conditions. The present study has shown that various concentrations of plant growth regulator can be effective for in vitro plant regeneration of H. syriacus. The direct and indirect regeneration protocol presented here will be useful for understanding the manipulation and propagation of H. syriacus.

Improvement of Shoot Regeneration from Scutella-Derived Callus in Rice

  • Kim, Yong-Wook;Cho, Joon-Hyeong;Lee, Jang-Yong
    • 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.

Adventitious Shoots Regeneration from Seed Explants of Xanthoceras sorbifolium

  • Hyunseok Lee
    • Proceedings of the Plant Resources Society of Korea Conference
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    • 2020.12a
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    • pp.58-58
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    • 2020
  • Xanthoceras sorbifolium Bunge (yellowhorn) is a woody tree in the soapberry family, Sapindaceae, native to northern China. This species has been identified as a major woody bioenergy plant for bio-diesel production because of high oil content in seed. But the flowers do not bear fruit well while the many flowers blooming. This study was performed to regenerate in vitro plantlet using adventitious shoot formation. To establish the protocol of plant regeneration, adventitious shoots formation rate in the culture of cotyledon of immature zygotic embryos was 68.6% in 1/2 MS medium with 0.5 mg l-1 BA and 3% sucrose (w/v). In the culture of cotyledons of mature zygotic embryos, induction of adventitious shoots was needed to contain high sucrose in pre-culture medium and the frequency of shoot induction was 64.4%. Multiple shoots were induced in 0.5 mg l-1 TDZ, and rooting of shoot was induced 4.0 mg l-1 IBA. Flow cytometry analysis revealed that all the regenerated plantlets were diploid.

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High-frequency regeneration by stem disc culture in selected clones of Populus euramericana

  • Cui, Hae-Yeon;Lee, Hyo-Shin;Oh, Chang-Young;Han, Shim-Hee;Lee, Kyung-Ju;Lee, Hyun-Jeong;Kang, Kyu-Seok;Park, So-Young
    • Journal of Plant Biotechnology
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    • v.41 no.4
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    • pp.236-241
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    • 2014
  • An efficient regeneration protocol for stem disc culture of Populus euramericana, which is important species for bioenergy resource in agroforestry, was established. The number of explants that were obtained and the number of explants that regenerated varied with the genotypes. However, in all the genotypes, stem disc culture produced more regenerated shoots than did in axillary bud culture. A comparison of the effects of cytokinin type and concentration on shoot regeneration in different explants (i.e., petiole, leaf, and root segments of P. euramericana) revealed that a concentration of $0.002mg\;l^{-1}$ thidiazuron (TDZ) used on petiole segments resulted in the greatest shoot regeneration (95.83%). The hormonal requirements for the greatest shoot regeneration in the three explant types varied. Different concentrations of $AgNO_3$ and $CoCl_2$ were added separately to the medium to stop the yellowing and subsequent necrosis of the regenerated shoots. Lower concentrations (3 and $5mg\;l^{-1}$) of these compounds improved shoot regeneration and elongation, compared with the control. The in vitro-regenerated shoots were transferred to rooting medium and subsequently acclimatized. The highly efficient regeneration system of P. euramericana reported here can be used for mass propagation of this recalcitrant for regeneration, economically important tree species.

In vitro Propagation and Ex vitro Rooting of Tectona grandis (L.f ), APNBV-1 Clone

  • Ramesh, Kommalapati;Chandra, Mouli Kalla;Vijaya, Tartte
    • Journal of Forest and Environmental Science
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    • v.25 no.2
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    • pp.119-126
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    • 2009
  • An efficient in vitro plant regeneration system was developed through shoot proliferation from axillary buds of Tectona grandis (L.f), APNBV-1 (Andhra Pradesh North Badrachalam Venkatapuram-1) clone. Multiple shoots of high quality were produced in vitro from axillary bud explants. An average of 4.39 shoots/explant were obtained on Murashige and Skoog's (MS) medium supplemented with plant growth regulators (PGRs) benzyl amino purine (BA), kinetin (KN), indole acetic acid (IAA), gibberillic acid ($GA_3$), growth adjuvants casein hydrolysate (CH), adenine sulphate (Ads) and antioxidants ascorbic acid, polyvinyl pyrrollidine (PVP). Eighty five percent of rooting was observed in ex vitro rooting media containing IBA and vermiculite. In ex vitro rooting, single shoots with 2 to 3 nodes were subjected to IBA of different concentrations at different periods of time intervals. Direct rooting in vermiculite at 500 ppm concentration of IBA resulted in 4.3 number of roots with 2 cm length. Minimum response of rooting and length of roots were recorded at 100 ppm concentration of IBA. Planlets were transferred to plastic bags for short acclimatization stage in green house where they survived at 95%.

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Efficient Callus Induction and Plant Regeneration from Immature and Mature Embryo Culture of Korean Wheat Genotypes

  • Lee, Byung-Moo;Moon, Jung-Hun;Lee, Sang-Kyu;Kim, Kyung-Hee;Kang, Moon-Seok;Heo, Hwa-Young;Kwon, Young-Up;Nam, Jung-Hyun;Seo, Yong-Weon
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.48 no.1
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    • pp.38-43
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    • 2003
  • Immature and mature embryos of 18 Korean wheat genotypes were cultured in vitro to develop an efficient method of callus formation and plant regeneration, and to compare the responses of both embryo cultures. Immature and mature embryos were placed on a solid agar medium containing the MS salts and vitamins, 30g/l maltose, 2 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D), and amino acids. The developed calli were maintained on regeneration medium containing MS salts and B5 vitamins, 20 g/l sucrose, and the combination of two plant growth regulators, 6-benzylaminopurine (BAP) and indole-3-acetic acid (IAA). Immature embryos in most genotypes showed high efficiency of callus induction except three genotypes; Eunpamil, Chunggemil, and Namhaemil, and significant differences among the genotypes. Plant regeneration of calli induced from immature embryos showed high efficiency in Geurumil (56.5%), Tapdongmil (50.5%), Gobunmil (45.5%), and Urimil(42.2%). The analysis of variance showed significant differences for regeneration frequency among the genotypes. Mature embryos showed low callus induction frequency compared with that in immature embryos, and significant differences among the genotypes. Plant regeneration of calli induced from mature embryos showed high efficiency in Keumkangmil (33.33%), Tapdongmil(28.13%), and Geurumil (27.78%). The analysis of variance showed significant differences for plant regeneration frequency among the genotypes.