• Journal of Plant Biotechnology
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• 제46권2호
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• pp.114-118
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• 2019

Efficient protocol for plant shoot regeneration of Brassica juncea L. CZERN was established by using organic media components and growth stimulating factors of the vermicompost and coelomic fluids. Formulated organic plant tissue culture media (Vermicompost (30%) extracts supplemented with 20 mL/L coelomic fluid) have shown maximum shoot regeneration when compared with the Murashige and Skoog (MS) medium, which were supplemented with 1 mg/L 6-benzyladenine (BA) and 0.1 mg/L of Naphthaleneacetic acid (NAA). Cotyledon explants produced the highest shoot regeneration frequency from fourday-old germinated seedlings in comparison with non-germinated seedlings. The vermicompost extracts have proved to be the best organic plant growth media to induce shoots from cotyledons compared to the MS media. Statistically significant difference (P = 0.008) for the root length, shoot length (P=0.000350) and the leaves (P=0.375) of the mustard plantlets were analyzed successfully. The survival rate was 98% in the mustard cotyledons on the Vermicompost extract media and 63% on MS media respectively. The coelomic fluid also is much suitable to induce shoots from cotyledons at lower concentrations. It was also shown that the vermicompost extract, which comprised of humic acids along with coelomic fluid, affected shoot regeneration from the cotyledons. An efficient and organic shoot regeneration study was standardized and it can be applicable in the improvement of the economically important crops.

• Journal of Plant Biotechnology
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• 제7권4호
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• pp.247-257
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• 2005

An efficient, rapid and large-scale in vitro clonal propagation of agronomically important Indian cereal crop genotypes (NSH27 & K5) of Sorghum bicolor (L.) Moench. by enhanced shoot proliferation in shoot tip segments was designed. MS medium fortified with plant growth regulators and coconut water markedly influenced in vitro propagation of Sorghum bicolor. In vitro plantlet production system has been investigated on Murashige and Skoog (MS) medium with the synergistic combination of 6-benzyladenine ($22.2\;{\mu}M$), kinetin ($4.6\;{\mu}M$), adenine sulphate ($2.8\;{\mu}M$), 5% coconut water and 3% sucrose which promoted the maximum number of shoots as well as beneficial shoot length. Subculturing of shoot tip segments on a similar medium enabled continuous production of more than 100 healthy shoots with similar frequency. When the healthy shoot clumps were cultured on MS medium fortified with 6-benzyladenine ($22.2\;{\mu}M$), kinetin ($4.6\;{\mu}M$), adenine sulphate ($2.8\;{\mu}M$), ${\alpha}$-naphthaleneacetic acid ($2.7\;{\mu}M$), ascorbic acid ($30.0\;{\mu}M$) and 5% coconut water, a rapid production of axillary and adventitious buds was developed after 8 wk culture. More than 300 shoots were produced 10 wk after culture. Rooting was highest (100%) on half strength MS medium containing 22.8 mM IAA. Micropropagated plants established in garden soil, farmyard soil and sand (2:1:1) were uniform and identical to the donor plant with respect to growth characteristics. These plants grew normally without showing any traits.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2018년도 추계학술대회
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• pp.89-89
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• 2018

우리나라, 중국 및 일본을 포함한 동북아시아가 원산이며 바이오매스량이 많은 억새(Miscanthus spp.)는 바이오에너지 생산을 위한 원료작물로서 가치가 높아, 바이오에탄올 생산용 원료작물로 주목을 받고 있다. 독일 등 유럽과 미국에서는 바이오에탄올 생산용 작물로 주로 종간 교잡 이질 3배체인 불임성 억새(M. x giganteous)를 대상으로 연구하고 있다. 이렇게 단일유전형을 갖는 품종의 재배에는 특정 병과 해충에 약하며 자연재해에도 취약성을 나타내므로 억새가 바이오에너지작물로 자리 잡기 위해서는 다양한 유전형의 억새 품종 개발이 필요하다. 본 연구는 우리나라의 자생 억새 3종을 기내배양하고 탈분화 및 재분화 시스템을 구축하여 억새 품종 육성 시 효율을 높이기 위해서 실시하였다. 억새 종자로부터 캘러스의 유도는 MS배지보다 N6배지 에서 좋았으며, 식물생장조절제로 2,4-Dichlorophenoxyacetic acid (2,4-D)와 6-Benzyl aminopurine (BA)를 조합처리한 처리구보다 2,4-D만을 단독처리하였을 때 캘러스 유도율이 더 높았다. 억새 종에 따른 캘러스 유도율은 물억새가 가장 낮고, 거대억새가 가장 높았으며, 3 ~ 5 mg/L의 2,4-D가 첨가된 N6배지에서 배발생 캘러스(embryogenic callus)가 발생하였다. 억새 신초 및 줄기의 절간에서의 캘러스 유도율은 전반적으로 종자에 비하여 낮았으며, 미성숙화기로부터의 캘러스 유도는 억새 종에 따른 차이가 없었으며, 5mg/L의 2,4-D가 첨가된 배지에서 캘러스 유도율이 가장 높게(90 ~ 95%) 나타났다. 형성된 배발생 캘러스로부터 식물체의 재분화는 N6배지에서는 재분화 식물체가 발생하지 않았고, 1 ~ 3mg/L의 BA와 0.1ml/L의 1-Naphthaleneacetic acid (NAA)가 첨가된 MS배지에서만 식물체가 재분화되였다.

• 한국응용곤충학회지
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• 제19권2호
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• pp.91-95
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• 1980

마늘의 바이러스 무감염주 생산과 종자 비용절감을 위한 기초적연구를 행하고저 마늘인편의 보통엽조직의 callus배양을 행하여 다음과 같은 결과를 얻었다. 1. 마늘 callus의 유도 Linsmaier & Skoog는 기본배지에 Benzyladenine $10^{-5}M$과 $2,4-D\;10^{-5}M$에서 가장 양호한 결과를 얻었다. 2. callus생장은 Linsmaier & Skoog 기본배지에 kinetin $10^{-6}M$과, $2,4D\;10^{-6}M$을 함유한 것이 가장 양호하였다. 3. callus조직내의 바이러스 소장을 조사한 결과 투명하고 부드러운 callus조직을 8대 계대배양을 하였을 때 바이러스는 제거되었다. 4. 바이러스무감염이 확인된 마늘 callus조직을 Murashing & Skoog 기본배지에 kinetin $10^{-5}M$와 $NAA\;5\times10^{-6}M$을 함유한 배지에 다식하였을 때 재분화 되어 소식물체를 형성하였다.

• Journal of Plant Biotechnology
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• 제34권4호
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• pp.293-298
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• 2007

Robinia pseudoacacia var. umbraculifera, commonly called umbrella black locust were regenerated after co-cultivation of internode segments with Agrobacterium tumefaciens which included yeast cadmium factor 1 (YCF 1) gene. The tolerance to cadmium and lead for plants can be increased by the YCF1 gene expression. Moreover, the recent studies have shown that YCF1 gene transgenic plants increase the accumulation of cadmium and lead into plant vacuoles. The effect of plant growth regulator such as 2,4-dichlorophenoxyacetic acid (2,4-D), ${\alpha}$-naphthaleneacetic acid (NAA), 6-benzyladenine (BA), and thidiazuron (TDZ) were studied to evaluate the propagation of plants through internode explants. The efficient induction of multiple adventitious shoots and callus were observed on a medium supplemented with 0.1 mg/L TDZ + 0.2 mg/L BA. To induce shoot elongation and rooting, regenerated shoots were transferred into basal MS medium without any plant growth regulator. Successful Agrobacterium tumefaciens mediated transformation was obtained by 20 min vacuum-infiltration with $50{\mu}M$ acetosyringone on the optimal multiple shoot induction medium with 30 mg/L hygromycin and 300 mg/L cefotaxime. To confirm the integration and expression of transgene, Polymerase Chain Reaction (PCR) and Reverse Transcriptase PCR (RT-PCR) were performed with specific primers. The frequency of transformation was approximately 18.94%. This study can be used to genetic engineering of phytoremediator.

• Journal of Plant Biotechnology
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• 제6권3호
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• pp.171-179
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• 2004

Hormonal requirements inducing different regeneration pathways with particular emphasis on somatic embryo-genesis in Alhagi graecorum were studied. While combination of 0.5 $\mu{M}$ 2,4-dichlorophenoxyacetic acid (2,4-D), 2.5 $\mu{M}$ 6-benzylaminopurine (BAP) and 5 $\mu{M}$ 1-naphthaleneacetic acid (NAA) in MS medium induced callus formation and callus maintenance from internodal explants, each alone or in combination with other induced distinct regeneration pathway. Adventitious bud formation was induced on MS medium supplemented with 2.5 $\mu{M}$ BAP. It was improved when 2.5 $\mu{M}$ BAP was used in combination with 5 $\mu{M}$ NAA. MS medium containing 0.5 $\mu{M}$ 2,4-D or 5 $\mu{M}$ NAA induced the formation of abnormal direct somatic embryos. While increase of 2,4-D concentration (1.125-9) resulted in the formation of viable embryogenic mass, increase of NAA did not change its effect. NAA should be used in combination with 2,4-D even at low concentration (0.5 $\mu{M}$) to form embryogenic mass. In A. gaecorum, the role of 2,4-D as trigger of somatic embryogenesis and BAP as trigger of adventitious bud formation was deduced, but for maximum yield certain auxin-cytokinin ratio should be applied. Embryogenic masses characterized by high water content, low peroxidase activity, and low number of peroxidase and glutamate oxaloacetate transaminase bands in comparison with calli obtained under conditions stimulating adventitious bud formation. The resulted differential gene expression, which could be detected by native-PAGE patterns, could be used as marker for organogenic pathway in A. graecorum.

• Journal of Plant Biotechnology
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• 제45권4호
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• pp.347-356
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• 2018

Chionanthus retusus is a small deciduous tree that is widely used in landscaping due to its beautiful white spring flowers and ornamental value. Conventional propagation through seeds requires one to two years of breaking dormancy. The objective of this study was to determine the conditions of in vitro germination in C. retusus. In vitro embryo culture was carried out to investigate the effects of six factors: basal media (McCown Woody Plant Medium (WPM) and Murashige and Skoog (MS)); plant growth regulators (different combinations and concentrations of naphthaleneacetic acid (NAA), 6-Benzylaminopurine (BA), and gibberellic acid ($GA_3$)); embryo age (collected weekly beginning 36 days after fruit setting); low temperature pretreatment (storing $4^{\circ}C$ for 1, 2, 3, and 4 weeks); coconut additives (100, 200, and $300ml{\cdot}L^{-1}$); and genotype (grouping plants depending on their flowering nature). The basal medium used in this study was WPM with $2mg{\cdot}L^{-1-1}\;GA_3$, $20g{\cdot}L^{-1}$ sucrose, and $6g{\cdot}L^{-1}$ Agar. WPM medium mixed with $GA_3$, resulted in higher germination rate as compared to when using a combination of auxin and cytokinin. $GA_3$ at $2mg{\cdot}L^{-1}$ was the most effective of all combinations and concentrations of PGRs. WPM medium with $2mg{\cdot}L^{-1}GA_3$ resulted in better and faster germination (75.93%). Embryos collected at 57 days after fruit setting had the highest percent of germinated seeds (87.04%) while low-temperature pretreatment of fruits at $4^{\circ}C$ for two weeks produced the highest germination (95.37%). These results of this study could be an open ground for development of an efficient protocol for commercial production of the ornamental tree.

• Journal of Plant Biotechnology
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• 제48권1호
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• pp.54-61
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• 2021

This work describe an efficient method for the shoot induction and plant regeneration of seedling-derived apical bud explants of Tilia mandshurica Rupr. & Maxim. The highest rate of shoot induction (82.2%) was obtained when apical bud explants from juvenile seedlings (5 months old) were cultured on Murashige and Skoog (MS) medium containing 1.0 mg/L 6-benzylaminopurine (BAP). However, apical bud explants obtained from mature trees (12 years old) did not produce any shoots, even with BAP supplementation. Among the three cytokinins tested for shoot multiplication (BAP, zeatin, and kinetin), BAP was the most effective; the highest number of shoots per explant (2.1) was observed on MS medium supplemented with 1.0 mg/L BAP. In contrast, the longest average shoot length (3.0 cm) was observed after growth on MS medium with 2.0 mg/L zeatin. No multiplication occurred when apical bud explants were cultured with kinetin-supplemented media. During rooting of in vitro-elongated shoots, the highest rooting rate (100%) was observed in half-strength MS medium supplemented with 0.5 ~ 1.0 mg/L indole-3-butyric acid (IBA) or 3.0 mg/L 1-naphthaleneacetic acid (NAA). During the acclimatization process, plantlets that were rooted on the IBA (0.5 mg/L)-supplemented medium had the highest survival rate (100%) and maximum root length (18.5 cm). These findings suggest that a low concentration (0.5 mg/L) of IBA is appropriate for the rooting and acclimatization of T. mandshurica. Plants were successfully transferred to the greenhouse with a 100% survival rate. This protocol will be useful for the large-scale propagation of Tilia species.

• Journal of Plant Biotechnology
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• 제45권4호
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• pp.382-391
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• 2018

본 연구는 사과 대목 M.26 (Malus pumila Mill))의 효과적인 기내 대량번식하기 위해 신초 증식과 뿌리 형성에 적합한 배지조건을 확립하고, simple sequence repeat (SSR) 마커를 이용하여 증식된 소식물체의 유전적 다양성을 분석하고자 수행하였다. MS (Murashige and Skoog) 기본배지에 benzyladenin (BA, $0.5{\sim}5.0mg{\cdot}L^{-1}$)와 thidiazuron (TDZ, $0.01{\sim}0.1mg{\cdot}L^{-1}$)을 첨가하여 신초를 배양한 결과, $1.0mg{\cdot}L^{-1}$ BA 처리에서 절편체 당신초 수가 10.67개로 가장 많았으며 과수화 발생률은 BA 처리구보다 TDZ 처리구에서 높았다. M.26 신초 증식에 BA와 auxin과의 혼용처리 효과는 없었고, $1.0mg{\cdot}L^{-1}$ BA가 첨가된 MS 기본배지가 적합하였다. 신초 발근에 적합한 배지를 구명하고자 auxin인 indole-3-butyric acid (IBA)와 ${\alpha}$-naphthaleneacetic acid의 농도($0.5{\sim}5.0mg{\cdot}L^{-1}$), MS 배지의 무기염류(1/4 ~ 1배) 및 sucrose 농도($0{\sim}30g{\cdot}L^{-1}$)를 달리하여 처리한 결과, $1.0mg{\cdot}L^{-1}$ IBA, $15{\sim}20g{\cdot}L^{-1}$ sucrose가 첨가된 1/2 MS 배지에서 발근율(100%), 뿌리 수(10.45 ~ 13.60개/절편체), 뿌리 길이(7.41 ~ 8.33 cm) 및 신초 길이(4.93 ~ 5.38 cm)가 양호하였다. 15종의 SSR primer를 이용하여 증식된 20개의 소식물체를 분석한 결과, 총 30개의 대립유전자가 검출되었으며 모두 동일한 밴드 패턴을 보여 온실에서 자란 M.26 식물체와 유사하여 유전적으로 안정한 것으로 판단되었다.

• Journal of Plant Biotechnology
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• 제47권1호
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• pp.53-65
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• 2020

Here, we describe an efficient and rapid protocol for the micropropagation of Thymus pallidus Cosson ex Batt., a very rare medicinal and aromatic plant in Morocco. After seed germination, we tested the effect of different macronutrients, cytokinins alone or in combination with gibberellic acid (GA₃) or auxins, on T. pallidus plantlet growth. We found that Margara macronutrients (N₃₀K) had the best effect on the in vitro development of the plantlets. The addition of 0.93 μM/L 1,3-diphenylurea (DPU), 0.46 μM/L adenine (Ad), and 0.46 and 0.93 μM/L kinetin (Kin) resulted in the best shoot multiplication and elongation. In addition, the combination of 0.46 μM/L Kin, DPU, or Ad with gibberellic acid, in particular, 0.46 μM/L Ad + 0.58 μM/L GA₃ and 0.46 μM/L Kin + 1.15 μM/L GA₃, led to better bud and shoot multiplication. Moreover, the integration of the combinations of 0.46 μM/L Kin and auxins, namely 0.46 μM/L Kin + 2.85 μM/L indole-3-acetic acid (IAA), 0.46 μM/L Kin + 2.85 or 5.71 μM/L indole-3-butyric acid (IBA), and 0.46 μM/L Kin + 0.3 or 0.57 μM/L 1-naphthaleneacetic acid (NAA), in the culture medium led to better root development and optimized aerial growth. Finally, the in vitro plants from the medium containing N₃₀K + 0.46 μM/L Kin + 2.85 μM/L IAA were successfully acclimatized; these plants served as a source for repeating in vitro culture.