• Proceedings of the KOSOMBE Conference
• /
• v.1997 no.11
• /
• pp.475-477
• /
• 1997

We have analyzed the fluorescence image obtaining from green fluorescence protein (GFP). In order to monitor the fluorescence of specific gene, we used the amyloid precursor protein promoter which has been known to act as a major role in the development of Alzheimer's disease. The promoter from - 3.0 kb to + 100 base pair was inserted into the gene expression monitoring GFP vector purchased from Clontech. This construct was transfected into the PC 12 and fibroblast cells and the fluorescence image was captured by two kinds of methods. One is using cheaper CCD camera and other is SIT-CCD camera. or the higher sensitivity of the fluorescence image, we developed the multiple image grabbing program. As a results, the fluorescence image by conventional CCD camera have the similar sensitivity compared with that of the SIT-camera by applying the multiple image grabbing programs. By this system. it will be possible to construct the fluorescence monitoring system with lower cost. And gene expression in real time by fluorescence image will be possible without changing the fluorescence images.

• Research in Plant Disease
• /
• v.15 no.3
• /
• pp.262-265
• /
• 2009

The fungus gnat, Bradysia difformis was examined for its ability to transmit Fusarium oxysporum in PDA culture. Larvae and adults of B. difformis were able to transmit the fungus as ingested and sticking. We constructed GFP-expressed mutants with Fusarium oxysporum, then feed it to larvae of fungus gnat, B. difformis. So that mycelia were placed in the alimentary canal of larva.

• Molecules and Cells
• /
• v.21 no.3
• /
• pp.401-410
• /
• 2006

The plastid transformation approach offers a number of unique advantages, including high-level transgene expression, multi-gene engineering, transgene containment, and a lack of gene silencing and position effects. The extension of plastid transformation technology to monocotyledonous cereal crops, including rice, bears great promise for the improvement of agronomic traits, and the efficient production of pharmaceutical or nutritional enhancement. Here, we report a promising step towards stable plastid transformation in rice. We produced fertile transplastomic rice plants and demonstrated transmission of the plastidexpressed green fluorescent protein (GFP) and aminoglycoside 3′-adenylyltransferase genes to the progeny of these plants. Transgenic chloroplasts were determined to have stably expressed the GFP, which was confirmed by both confocal microscopy and Western blot analyses. Although the produced rice plastid transformants were found to be heteroplastomic, and the transformation efficiency requires further improvement, this study has established a variety of parameters for the use of plastid transformation technology in cereal crops.

• Molecules and Cells
• /
• v.38 no.11
• /
• pp.959-965
• /
• 2015

Inducible and reversible gene silencing in desired types of cells is instrumental for deciphering gene functions using cultured cells or in vivo models. However, efficient conditional gene knockdown systems remain to be established. Here, we report the generation of an inducible expression system for short hairpin RNA (shRNA) targeted to PTEN, a well-documented dual-specificity phosphatase involved in tumor suppression and ontogenesis. Upon induction by doxycycline (DOX), the reverse tetracycline transcriptional activator (rtTA) switched on the concomitant expression of GFP and a miR-30 precursor, the subsequent processing of which released the embedded PTEN-targeted shRNA. The efficacy and reversibility of PTEN knockdown by this construct was validated in normal and neoplastic cells, in which PTEN deficiency resulted in accelerated cell proliferation, suppressed apoptosis, and increased invasiveness. Transgenic mice harboring the conditional shRNA-expression cassette were obtained; GFP expression and concurrent PTEN silencing were observed upon ectopic expression of rtTA and induction with Dox. Therefore, this study provides novel tools for the precise dissection of PTEN functions and the generation of PTEN loss of function models in specific subsets of cells during carcinogenesis and ontogenesis.

• Journal of Microbiology and Biotechnology
• /
• v.24 no.4
• /
• pp.568-576
• /
• 2014

TIM-1 (also known as KIM-1 and HAVcr-1) is a type I transmembrane glycoprotein member of the TIM family that may play important roles in innate and adaptive immune responses. The overexpression of proteins associated with membrane proteins is a major obstacle to overcome in studies of membrane protein structures and functions. In this study, we successfully coupled the overexpression of the TIM-1 protein with a C-terminal enhanced green fluorescent protein (GFP) tag in Escherichia coli. To the best of our knowledge, this report is the first to describe the overexpression of human TIM-1 in E. coli. The purified TIM-1-EGFP fusion protein recognized and bound directly to apoptotic cells and did not to bind to viable cells. Furthermore, we confirmed that the interactions of TIM-1-EGFP with apoptotic cells were blocked by TIM-1-Fc fusion proteins. This fusion protein represents a readily obtainable source of biologically active TIM-1 that may prove useful in future studies of human TIM-1.

• Journal of Life Science
• /
• v.21 no.1
• /
• pp.159-164
• /
• 2011

Microbial whole-cell biosensors can be excellent analytical tools for monitoring environmental pollutants. They are constructed by fusing reporter genes (e.g., lux, gfp or lacZ) to inducible regulatory genes which are responsive to the relevant pollutants, such as aromatic hydrocarbons and heavy metals. A large spectrum of microbial biosensors has been developed using recombinant DNA technology and applied in fields as diverse as environmental monitoring, medicine, food processing, agriculture, and defense. Furthermore, their sensitivity and target range could be improved by modification of regulatory genes. Recently, microbial biosensor cells have been immobilized on chips, optic fibers, and other platforms of high-throughput cell arrays. This paper reviews recent advances and future trends of genetically modified microbial biosensors used for monitoring of specific environmental pollutants.

• Molecules and Cells
• /
• v.37 no.5
• /
• pp.399-405
• /
• 2014

Autophagy targets cytoplasmic cargo to a lytic compartment for degradation. Autophagy-related (Atg) proteins, including the transmembrane protein Atg9, are involved in different steps of autophagy in yeast and mammalian cells. Functional classification of core Atg proteins in plants has not been clearly confirmed, partly because of the limited availability of reliable assays for monitoring autophagic flux. By using proUBQ10-GFP-ATG8a as an autophagic marker, we showed that autophagic flux is reduced but not completely compromised in Arabidopsis thaliana atg9 mutants. In contrast, we confirmed full inhibition of auto-phagic flux in atg7 and that the difference in autophagy was consistent with the differences in mutant phenotypes such as hypersensitivity to nutrient stress and selective autophagy. Autophagic flux is also reduced by an inhibitor of phosphatidylinositol kinase. Our data indicated that atg9 is phenotypically distinct from atg7 and atg2 in Arabidopsis, and we proposed that ATG9 and phosphatidylinositol kinase activity contribute to efficient autophagy in Arabidopsis.

• Proceedings of the Korean Society of Embryo Transfer Conference
• /
• 1998.05a
• /
• pp.12-28
• /
• 1998

The technology of creating transgenic animals has a potential value in improving productivity and disease resistance of animals, gene therapy, drug pharming and production of model animals for certain diseases. Up to date, fairly low success rate of production of transgenic animals and a pronounced variability with respect to the expression of transgenes have been much observed. The mechanisms how to integrate the injected genes with a certain part of the genomes are unknown yet. Many techniques in gene transfer, beside microinjection, have been introduced and explored thus to improve the production efficiency of transgenic animals. In this article, the methods and efficiency of gene-transfer techniques, the detection and preselection of transgenes in embryos by PCR- and GFP-screenings and cloning of preselected transgenic embryos by nuclear transplantation are described and discussed. Some experimental results showed that the early screening and selection of integration of the injected gene with embryonic genome by polymerase chain reaction(PCR) and green fluorecence protein(GFP) were promising methods. Further, the application of nuclear transplantation technology to cloning and multiplication of the positively integrated genes in the cleaving embryos and embryonic cells will be beneficially used for the mass production of transgenic embryos and consequently improving the production efficiency in transgenic animals.

• Molecules and Cells
• /
• v.38 no.1
• /
• pp.51-57
• /
• 2015

The Caenorhabditis elegans peroxidasins, PXN-1 and PXN-2, are extracellular peroxidases; pxn-2 is involved in muscle-epidermal attachment during embryonic morphogenesis and in specific axon guidance. Here we investigate potential roles of the other homologue of peroxidasin, pxn-1, in C. elegans. A pxn-1 deletion mutant showed high lethality under heat-stress conditions. Using a transcriptional GFP reporter, pxn-1 expression was observed in various tissues including neurons, muscles, and hypodermis. A translational fusion showed that PXN-1::GFP was secreted and localized in extracellular matrix, particularly along body wall muscles and pharyngeal muscles. Various neuronal developmental defects were observed in pxn-1 mutants and in pxn-1 over-expressing animals, including handedness, branching, breakage, tangling, and defasciculation. These results suggest that pxn-1, like other peroxidasins, plays an important role throughout development.

• ALGAE
• /
• v.24 no.2
• /
• pp.85-91
• /
• 2009

The filamentous cyanobacterium Anabaena sp. Strain PCC 7120 produces a developmental patten of single hete- rocysts separated by approximately 10 vegetative cells. Heterocysts differentiate from vegetative cells and are spe- cialized for nitrogen fixation. The patS gene, which encodes a small peptide that inhibits heterocyst differentiation, is expressed in proheterocysts and plays a critical role in establishing the heterocyst pattem. Another key regulator of heterocyst development is the hetR gene. hetR mutants fail to produce heterocysts and extra copies of hetR on a plas- mid cause a multiple contiguous heterocyst phenotype. To elucidate the relationship between these two counter act- ing factors in the genetic regulatory pathway during heterocyst differentiation, the expression patterns of a patS-gfp and a hetR-gfp fusion were examined in a patS deletion and a hetR deletion strain. The results, in combination with the result from a hetR and patS double deletion strain, suggest patS and hetR are mutually antagonistic and the bal- ance between these two factors in tow different cell types (heterocysts and vegetative cells) may be critical during the decision making process on their cell fates.