• Proceedings of the Plant Resources Society of Korea Conference
• /
• 2003.04a
• /
• pp.61-62
• /
• 2003

Clonal propagation of high-value forest trees through somatic embryogenesis (SE) has the potential to rapidly capture the benefits of breeding or genetic engineering programs and to improve raw material uniformity and quality. A major barrier to the commercialization of this technology is the low quality of the resulting embryos. Several factors limit commercialization of SE for Corsican pine, including low initiation rates, low culture survival, culture decline causing low or no embryo production, and inability of somatic embryos to fully mature, resulting in low germination and reduced vigour of somatic seedlings. The objective was to develop a Corsican pine maturation medium that would produce cotyledonary embryos capable of germination. Treatments were arranged in a completely randomized design. Data were analyzed by analysis of variance, and significant differences between treatments determined by multiple range test at P=0.05. Corsican pine (Pinus nigra var. maritima) cultures were initiated on modified !P6 medium. Modifications of the same media were used for culture multiplication and maintenance. Embryogenic cultures were maintained on the same medium semi solidified with 2.5 g/l Gelrite. A maturation medium, capable of promoting the development of Corsican pine somatic embryos that can germinate, is a combination of iP6 modified salts, 2% maltose, 13% polyethylene glycol (PEG), 5 mg!l abscisic acid (ABA), and 2.5 g/l Gelrite. After initiation and once enough tissue developed they were grown in liquid medium. Embryogenic cell suspensions were established by adding 0.951.05 g of 10- to 14-day-old semisolid-grown embryogenic tissue to 9 ml of liquid maintenance media in a 250ml Erlenmeyer flask. Cultures were then incubated in the dark at 2022$^{\circ}$C and rotated at 120 rpm. After 2.53 months on maturation medium, somatic embryos were selected that exhibited normal embryo shape. Ten embryos were placed horizontally on 20 ml of either germination medium ($\frac{2}{1}$strength Murashige and Skoog (1962) salts with 2.5 g/l activated charcoal) or same medium with copper sulphate adjusted to 0.25 mg/1 to compensate for copper adsorption by activated carbon. 2% and 4% maltose was substituted by 7.5% and 13% PEG respectively to improve the yield of the embryos. Substitution of' maltose with PEG was clearly beneficial to embryo development. When 2% of the maltose was replaced with 7.5% PEG, many embryos developed to large bullet-shaped embryos. At latter stages of development most embryos callused and stopped development. A few short, barrel-shaped cotyledonary embryos formed that were covered by callus on the sides and base. When 4% of the maltose was removed and substituted with 13% PEG, the embryos developed further, emerging from the callus and increasing yield slightly. Microscopic examination of the cultures showed differing morphologies, varying from mostly single cells or clumps to well-formed somatic embryos that resembled early zygotic embryos only liquid cultures with organized early-stag. A procedure for converting and acclimating germinants to growth in soil and greenhouse conditions is also tested. Seedling conversion and growth were highly related to the quality of the germinant at the time of planting. Germinants with larger shoots, longer, straighter hypocotyls and longer roots performed best. When mature zygotic embryos germinate the root emerges, before or coincident with the shoot. In contrast, somatic embryos germinate in reverse sequence, with the cotyledons greening first, then shoot emergence and then, much later, if at all, the appearance of the root. Somatic seedlings, produced from the maturation medium, showed 100% survival when planted in a field setting. Somatic seedlings showed normal yearly growth relative to standard seedlings from natural seed.

• Korean Journal of Plant Resources
• /
• v.34 no.1
• /
• pp.59-63
• /
• 2021

To study for germination rate of apple artificial seeds encapsulated with 2% alginic acids, the rates of germination for artificial seeds of various embryos with partial elimination of cotyledon measured by adding 80 mg/L gibberellic acid or/and nutritional media. The germination rate of artificial seeds produced embryo with 3/4 cotyledon (3/4 CE), embryo with 1/2 cotyledon (1/2 CE), embryo with 1/4 cotyledon (1/4 CE), and embryo without cotyledon (NCE) were gradually decreased with compare to normal embryo, but it's considerably recovered on the alginic acid containing MS solution or/and 80 mg/L GA3. In special, the low frequency (55.81%) of germination for artificial seeds of 1/2 cotyledon embryo with alginic acid was increased by treatment with combination of alginic acid, 3% sucrose, and 80 mg/L gibberellic acid (98.74%). In case of embryo without cotyledon, the low frequency of embryo without cotyledon (24.44%) in alginic acid almost recovered to 82.45% in alginic acid with MS solution and 80 mg/L gibberellic acid. The results shown that the germination of embryo was decreased by the removal of cotyledon, and demonstrated that the low rate of germination could be considerably increased by adding GA3 (80 mg/L) or/and nutrition.

• Korean Journal of Plant Tissue Culture
• /
• v.28 no.1
• /
• pp.21-24
• /
• 2001

Peucedanum japonicum $T_{HUNB}$ used as a edible and medicinal plants was investigated for in uitro regeneration. Callus formation occurred on leaf and stem explant cultures and showed spontaneous embryogenic and organogenic capability on MS basal medium supplemented with 0.1~5 mg/L NAA and 0~10 mg/L BA in dark. The regeneration was highest on the condition supplemented with 2.5 mg/L NAA and 10 mg/L BA. Development of the somatic embryo progressed through the globular, heart-shaped, torpedo-shaped and cotyledonary stage, typical of zygotic embryos. When the first somatic embryos was cultured on the medium supplemented with 0.2 mg/L NAA, secondary somatic embryo were induced with higher frequency on the hypocotyl then on the cotyledon and root.t.

• Korean Journal of Plant Tissue Culture
• /
• v.27 no.4
• /
• pp.245-258
• /
• 2000

Somatic embryogenesis has become a major tool in the study of plant embryology, as it is possible in culture to manipulate cells of many plant species to produce somatic embryos in a process that is remarkably similar to zygotic embryogenesis. Traditionally, the process has been studied by an examination of the ex vitro factors which influence embryo formation. Later structural, physiological and biochemical approaches have been applied. Host recently, molecular tools are being used. Together, these various approaches are giving valuable information on the process. This article gives an overview of somatic embryogenesis by reviewing information on the morphogenesis, physiology, biochemistry and molecular biology of the process. Topics covered include a brief description of the factors involved in the production of embryogenic cells. Carrot cell suspension is most commonly used, and the development of a high frequency and synchronous system is outlined. At the physiological and biochemical lev-els various topics, including the reactivation of the cell cycle, changes in endogenous growth regulators, amino acid, polyamine, DNA, RNA and protein metabolism, and embryogenic factors in conditioned medium are all discussed. Lastly, recent information on genes and molecular markers of the embryogenic process are outlined. Somatic embryogenesis, the best example of totipotency in plant cells, is not only an important tool in studies in basic biology, but is potentially of equal significance in the micropropagation of economically important plants.

• Development and Reproduction
• /
• v.23 no.3
• /
• pp.285-295
• /
• 2019

Junction-mediating and regulatory protein (JMY) is a regulator of both transcription and actin filament assembly. The actin-regulatory activity of JMY is based on a cluster of three actin-binding Wiskott-Aldrich syndrome protein homology 2 (WH2) domains that nucleate actin filaments directly and promote nucleation of the Arp2/3 complex. In addition to these activities, we examined the activity of JMY generation in early embryo of mice carrying mutations in the JMY gene by CRISPR/Cas9 mediated genome engineering. We demonstrated that JMY protein shuttled expression between the cytoplasm and the nucleus. Knockout of exon 2, CA (central domain and Arp2/3-binding acidic domain) and NLS-2 (nuclear localization signal domain) on the JMY gene by CRISPR/Cas9 system was effective and markedly impeded embryonic development. Additionally, it impaired transcription and zygotic genome activation (ZGA)-related genes. These results suggest that JMY acts as a transcription factor, which is essential for the early embryonic development in mice.

• Korean Journal of Medicinal Crop Science
• /
• v.8 no.1
• /
• pp.14-20
• /
• 2000

This experiment was conducted to investigate the effects of length of storage period under low temperature, $CO_2$ enrichment and addition of plant growth regulators in Murashige and Skoog medium on the plant regeneration of Korean ginseng (Panax ginseng C. A. Meyer). Seeds were treated for 60 and 80 days respectively under $5^{\circ}C$ environment. 2500ppm of $CO_2$ was enriched by ventilation. Three plant growth regulators added to the medium were Indolbutyric acid, Benzyladenin and Gibberellic acid (GA3). The result indicated that : The capacity of differentiation was higher in the aged cotyledons from the seeds treated for 80 days under low temperature condition than in those treated for 60 days. $CO_2$ enrichment had stimulating effects on the growth and development of shoot primordium significantly but less effects on the formation of adventitious buds. From one zygotic embryo hundreds of plantlets were differentiated. $CO_2$ enrichment had effects on the formation of both indirect somatic embryo and direct somatic embryo. Indirect somatic embryo showed little growth and differentiation, being undifferentiated vascular stele and epicotyl. Direct somatic embryos were formed on the epidermis of backside basal part of cotyledon. Those embryos developed to whole plant having latent bud.

• Journal of Plant Biotechnology
• /
• v.42 no.4
• /
• pp.388-395
• /
• 2015

Somatic embryogenesis is as an excellent technology for potential use in plant mass production, germplasm conservation, or genetic engineering. We examined the effect of cold storage using 3 embryogenic callus lines with different levels of embryogenesis competence derived from immature zygotic embryo cultures of Kalopanax setemlobus. Somatic embryo induction, germination and plant conversion were evaluated after 1, 3 and 6 months storage at $4^{\circ}C$ in the dark. Most cold-stored embryogenic calli formed somatic embryos normally even after 6 months; however, the induction rate was gradually decreased by increasing the storage period. The most competent line tended to show a slight decline in somatic embryo induction rate, as compared with other lines after cold storage. In general, cold storage resulted in reduced somatic embryo germination and plant regeneration, although 93% somatic embryo germination and 91% plant conversion were achieved regardless of the storage period. Cold storage led to cell browning and degradation. Additionally, the cell structures were confirmed by the aceto-carmine and evans blue dye evaluation. Collectively, our results showed that embryogenic callus of K. septemlobus could be preserved at $4^{\circ}C$ without subculture for 6 months, and suggested the need for storage of relatively more competent embryogenic calli lines to support somatic embryo induction.

• Journal of the Korea Academia-Industrial cooperation Society
• /
• v.12 no.8
• /
• pp.3541-3546
• /
• 2011

Autophagy is an evolutionarily conserved lysosomal pathway for degrading cytoplasmic proteins, macromolecules, and organelles in addition to recycling protein and ATP synthesis. Although autophagy is very important during embryogenesis, the mechanism underlying the dynamic development during this process remains largely unknown. In order to obtain insights into autophagy in early embryo development, we analyzed gene expression levels of autophagy-related genes (ATGs) in mouse embryos developing in vitro. Using real time RT-PCR technique, ATGs including Atg2a, Atg3, Atg4b, Atg5, Atg6, Atg7, Atg9a, and Wipi3, as maternal transcripts, were only up-regulated in 1-cell embryo stage before zygotic genomic activation (ZGA), and then expression decreased from 2-cell to blastocyst embryo stage. ATGs including Dram and Atg9b were expressed abundantly in 1-cell embryo state and in blastocyst embryo stage, athough Atg8 and Ulk1 were constantly expressed during preimplantation stage. However, Atg4d were only up-expressed from 4-cell to blastocyst stage. These results suggest that autophagy is related in mouse embryo, which possibly gives an important role for early development.

• Animal cells and systems
• /
• v.13 no.4
• /
• pp.411-418
• /
• 2009

We are reporting the identification, expression patterns, and possible biological functions of zebrafish kif3b (kif3bz) encoding 475 amino acids. Kif3Bz contains the kinesin motor domain, catalytic domain, KISc domain, and one single coiled coil domain. Phylogenetic analysis indicates that kif3bz is a highly conserved gene among the tested vertebrates. First of all, both maternal and zygotic messages of kif3bz were evenly distributed in the blastomeres at 2-cell stage. Its ubiquitous expression throughout the blastomeres continued till 40% epiboly. However, kif3bz transcripts became restricted in Kupffer's vesicle at tailbud and 6-somite stages. At 13-somite stage, kif3bz expression pattern became specific to the telencephalon, diencephalon, trigeminal placode, and somites. Such expression patterns were further intensified in the telencephalon, diencephalons, hind brain, pronephric ducts, optic vesicles, and spinal cord neurons in the 23-somite stage embryos, and last till 24 hpf. We discussed possible functions of Kif3Bz related to the vertebrate embryonic development.

• Journal of Plant Biotechnology
• /
• v.50
• /
• pp.267-274
• /
• 2023

All studies on date palm somatic embryogenesis have focused on germination in the presence of light while neglecting germination in darkness, which mimics the germination process of zygotic embryos within seeds. To improve the date palm micropropagation protocol, we investigated the effects of light and darkness incubation on the germination of indirect date palm somatic embryos and their subsequent conversion into plantlets. Darkness incubation emerged as a pivotal factor in the germination of indirect date palm somatic embryos and their successful conversion into plantlets. Darkness incubation significantly decreased the time required for the conversion of indirect somatic embryos into plantlets, halving the duration from 24 weeks to only 12 weeks. The micropropagation protocol was modified, consolidating the previous two distinct stages of germination and elongation under light incubation into a single stage under darkness incubation. These findings modified the protocol and significantly reduced the overall duration of the date palm micropropagation protocol.