• Journal of Life Science
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• v.14 no.6 s.67
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• pp.1018-1022
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• 2004

Optimized conditions for Agrobacterium-mediated lisianthus pollen transformation were adjusted using various factors such as temperature, pH and sucrose concentration. Pollen tube growth was successfully achieved in a medium (pollen germination medium; PGM) containing $7-15\%$ sucrose with pH in the range of 5.5-7.0 at temperature of $20-27^{\circ}C$. Lisianthus pollen was vacuum-infiltrated with Agrobacterium cell suspension for 20 min, and transformed pollen was confirmed by GUS histochemistry and Southern hybridization following RT-PCR. Transgenic pollen system may be utilized for establishing an area of plant transient expression systems based on the convenient pollen transformation procedure presented in here.

• Journal of agriculture & life science
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• v.44 no.1
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• pp.9-15
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• 2010

Barley, a monocotyledonous plant, is relatively recalcitrant to the process of Agrobacterium-mediated genetic transformation. In this study, seedlings of six barley cultivars (Keunal-1-Ho, Saessal, Ol, Saechalssal, Seodunchal and Pungsanchalssal) were injured using alkali, oxidizing or reducing agents. They were then transformed using Agrobacterium via vacuum infiltration for the analysis of comparative GUS gene expression. It was determined that chemical injuries causing a slight growth retardation could overall enhance the GUS transformation rate. Hydrogen peroxide was determined to be the most effective.

• Journal of Ginseng Research
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• v.41 no.3
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• pp.419-427
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• 2017

Background: Glycosylation of natural compounds increases the diversity of secondary metabolites. Glycosylation steps are implicated not only in plant growth and development, but also in plant defense responses. Although the activities of uridine-dependent glycosyltransferases (UGTs) have long been recognized, and genes encoding them in several higher plants have been identified, the specific functions of UGTs in planta remain largely unknown. Methods: Spatial and temporal patterns of gene expression were analyzed by quantitative reverse transcription (qRT)-polymerase chain reaction (PCR) and GUS histochemical assay. In planta transformation in heterologous Arabidopsis was generated by floral dipping using Agrobacterium tumefaciens (C58C1). Protein localization was analyzed by confocal microscopy via fluorescent protein tagging. Results: PgUGT72AL1 was highly expressed in the rhizome, upper root, and youngest leaf compared with the other organs. GUS staining of the promoter: GUS fusion revealed high expression in different organs, including axillary leaf branch. Overexpression of PgUGT72AL1 resulted in a fused organ in the axillary leaf branch. Conclusion: PgUGT72AL1, which is phylogenetically close to PgUGT71A27, is involved in the production of ginsenoside compound K. Considering that compound K is not reported in raw ginseng material, further characterization of this gene may shed light on the biological function of ginsenosides in ginseng plant growth and development. The organ fusion phenotype could be caused by the defective growth of cells in the boundary region, commonly regulated by phytohormones such as auxins or brassinosteroids, and requires further analysis.

• Journal of Radiation Industry
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• v.10 no.4
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• pp.243-248
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• 2016

Argonaute 2 (AtAGO2) is a well characterized effector protein in Arabidopsis for its functionalities associated with DNA double-strand break (DSB)-induced small RNAs (diRNAs) and for its inducible expression upon ${\gamma}$-irradiation. However, its transcriptional regulation depending on the recovery time after the irradiation and on the specific response to DSBs has been poorly understood. We analyzed the 1,313 bp promoter sequence of the AtAGO2 gene ($1.3kb_{pro}$) to characterize the transcriptional regulation of AtAGO2 at various recovery times after ${\gamma}$-irradiation. A stable transformant harboring $1.3kb_{pro}$ fused with GUS gene showed that the AtAGO2 is highly expressed in response to ${\gamma}$-irradiation, after which the expression of the gene is gradually decreased until 5 days of DNA damage recovery. We also confirm that the AtAGO2 expression patterns are similar to that of ${\gamma}$-irradiation after the treatments of radiomimetic genotoxins (bleomycin and zeocin). However, methyl methanesulfonate and mitomycin C, which are associated with the inhibition of DNA replication, do not induce the expression of the AtAGO2, suggesting that the expression of the AtAGO2 is closely related with DNA DSBs rather than DNA replication.

• Korean Journal of Plant Tissue Culture
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• v.25 no.4
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• pp.237-243
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• 1998

To understand the tissue specific expression pattern of S RNase genes associated with self-incompatibility in L. peruvianum, two promoter regions of $S_{11}$ and $S_{12}$ RNase genes were compared. Homologous sequences between two S RNase gene promoters were found within 300 bp upstream of transcription start site. Moreover short direct repeat sequences within $S_{11}$ RNase gene promoter existed in the vicinity of 350-500 bp upstream of transcription start site. To identify whether the unique promoter sequences of $S_{11}$ RNase gene confer the tissue specific expression, six deletion fragments for $S_{11}$ genomic gene promoter constructed by PCR were fused to $\beta$-glucuronidase gene, and introduced into various tissues of L. peruvianum by microprojectile bombardment. Transient expression assays indicated that $S_{11}$ RNase gene promoter contained the positive and negative regulatory sequences, which can control the floral tissue-specific expression in L. peruvianum.

• Journal of Plant Biotechnology
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• v.4 no.2
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• pp.71-76
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• 2002

In order to achieve repetitive somatic embryogenesis in cacao (Theobroma cacao L.), callus derived from floral tissues were continuously cultured in a medium containing 2,4-D. In 5% of the explants, repetitive somatic embryogenesis was observed after 8 weeks and maintained in a globular stage for several weeks. This is the first report showing repetitive somatic embryogenesis in cacao. The calli were bombarded with a plasmid containing $\beta$-glucuronidase (gus) as reporter gene. Two week old calli showed the high average number of cells expressing the us gene. The effect of osmotic agents (mannitol, sorbitol and sucrose) on gene expression was evaluated. Pre-treatment during 16 h with 0.25 M mannitol revealed an improvement in gene expression. The potential utilization of the repetitive embryogenesis, combined with osmotic treatment, is discussed as an alternative to achieve stable transgenic cacao plants.

• Journal of Plant Biotechnology
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• v.7 no.2
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• pp.113-121
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• 2005

The objective of this study was to identify the major parameters controlling DNA delivery by particle bombardment to immature embryos of Chinese spring wheat (Triticum aestivum L.). Efficiency of DNA (uidA gene) delivery was assessed by transient GUS (${\beta}$-glucuronidase) expression in bombarded tissues. Of the parameters analyzed, acceleration pressure, bombardment distance, chamber vacuum pressure, bombardment times, osmotic conditioning of culture had a remarkable influence on transient gene expression. A bombardment procedure suitable for Chinese spring wheat cultivars was developed which allowed high-efficiency DNA delivery combined with reduced damage to target tissues. The high efficiency made the system practical for wheat genetic transformation research and accelerating wheat breeding programs.

• Journal of Plant Biotechnology
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• v.50
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• pp.56-62
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• 2023

Transgenic lilies have been obtained using Agrobacterium tumefaciens (AGL1) with the plant scale explants, followed by DL-phosphinothricin (PPT) selection. In this study, scales of lily plants cv. "red flame" were transformed with the pCAMBIA3301 vector containing the gus gene as a reporter and the blpR gene as a selectable marker, as well as a gene of interest showing herbicide tolerance, both driven by the CaMV 35S promoter. Using a 20-minute infection time and a 5-day cultivation period, factors that optimized and demonstrated a high transformation efficiency were achieved. With these conditions, approximately 22-27% efficiency was observed for Agrobacterium-mediated transformation in lilies. After transformation with Agrobacterium, scales of lilies were transferred to MS medium without selective agents for 2 weeks. They were then placed on selection MS medium containing 5 mg/L PPT for a month of further selection and then cultured for another 4-8 weeks with a 4-week subculture regime on the same selection medium. PPT-resistant scales with shoots were successfully rooted and regenerated into plantlets after transferring into hormone-free MS medium. Also, most survived putatively transformed plantlets indicated the presence of the blpR gene by PCR analysis and showed a blue color indicating expression of the gus gene. In conclusion, when 100 scales of lily cv. "red flame" are transformed with Agrobacterium, approximately 22-27 transgenic plantlets can be produced following an optimized protocol. Therefore, this protocol can contribute to the lily breeding program in the future.

• Applied Biological Chemistry
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• v.39 no.4
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• pp.260-264
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• 1996

Process of particle bombardment for efficient transformation of Cymbidium virescence rhizome microcross sections was investigated using Biolistic particle delivery system with pBI121 harboring the ${\beta}-glucuronidase$(GUS) and the neomycin phosphotransferaseII(nptII). The best result was obtained from the combination of $1.11{\;}{\mu}m$ tungsten particles coated with pBl121, $77.33kg/cm^2$ helium pressure, 6.35 mm gap distance, and 7.0 cm target distance. Transient expression of the reporter gene, GUS, bombarded into the rhizome microsections was observed by the histochemical assay. The marker gene, nptII, delivered by bombarding the tungsten particles coated with the plasmid DNA was identified in the transformed rhizome by polymerase chain reaction.

• Journal of Plant Biotechnology
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• v.35 no.3
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• pp.177-183
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• 2008

Ac/Ds mutant lines of this study were transgenic rice plants, each of which harbored the maize transposable element Ds together with a GUS coding sequence under the control of a promoterless(Ds-GUS). We selected the mutants that were GUS expressed lines, because the GUS positive lines will be useful for identifying gene function in rice. One of these mutants was identified knock-out at Oszinc626(NP_001049991) gene, encoding a RING-H2 zinc-finger protein, by Ds insertion. In this mutant, while primary root development is normal, secondary root development from lateral root was very poor and seed development was incomplete compare with normal plant. RING zinc-finger proteins play important roles in the regulation of development in a variety of organisms. In the plant kingdom, a few genes encoding RING zinc-finger proteins have been documented with visible effects on plant growth and development. The consensus of the RING-H2(C3-H2-C3 type) domain for this group of protein is $Cys-X_2-Cys-X_{28}-Cys-X-His-X_2-His-X_2-Cys-X_{14}-Cys-X_2-Cys$. Oszinc626 encodes a predicted protein product of 445 amino acids residues with a molecular mass of 49 kDa, with a RING-zinc-finger motif located at the extreme end of the C-terminus. RT-PCR analysis indicated that the expression of Oszinc626 gene was induced by IAA, cold, dehydration, high-salinity and abscisic acid, but not by 2,4-D, and the transcription of Oszinc626 gene accumulated primarily in rice immature seeds, root meristem and shoots. The gene accumulation patterns were corresponded with GUS expression.