• 한국생물공학회:학술대회논문집
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• 2005.04a
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• pp.408-411
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• 2005

The green fluorescent protein (GFP) from the jellyfish aequorea and its fluorescent homologs from Anthozoa corals have become invaluable tools for imaging of cells and tissues. In this study various fluorescent protein such as green fluorescent protein (GFP), yellow fluorescent protein (YFP) and red fluorescent protein (RFP) have been expressed in Escherichia coli. Growth of recombinant cells and production of fluorescent proteins were investigated in shake flasks. Some characteristics of fluorescent proteins was also studied.

• International Journal of Industrial Entomology and Biomaterials
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• v.45 no.2
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• pp.56-69
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• 2022

Silk is a unique natural biopolymer with outstanding biocompatibility, high mechanical strength, and superior optical transparency. Due to its excellent properties, silk has been widely reported as an ideal biomaterial for several biomedical applications. Recently, fluorescent silk protein, a variant of native silk, has been reported as a biophotonic material with the potential for bioimaging and biosensing. Despite the realization of fluorescent silk, the traditional degumming process of fluorescence silk is crude and often results in fluorescence loss. The loss of fluorescent properties is attributed to the sensitivity of silk fibroin to temperature and solvent concentration during degumming. However, there is no comprehensive information on the influence of these processing parameters on fluorescence evolution and decay during fluorescent silk processing. Therefore, we conducted a spectroscopic study on fluorescence decay as a function of temperature, concentration, and duration for fluorescent silk cocoon degumming. Sodium carbonate solution was tested for degumming the fluorescent silk cocoons with different concentrations and temperatures; also, sodium carbonate solution is combined with Alcalase enzyme and triton x-100 to find optimal degumming conditions. Additionally, we conducted a molecular dynamics study to investigate the fundamental effect of temperature on the stability of the fluorescent protein. We observed degumming temperature as the prime source of fluorescent intensity reduction. From the MD study, fluorescence degradation originated from the thermal agitation of fluorescent protein Cα atoms and fluctuations of amino acid residues located in the chromophore region. Overall, degumming fluorescent silk with sodium carbonate and Alcalase enzyme solution at 25 ℃ preserved fluorescence.

• International Journal of Industrial Entomology and Biomaterials
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• v.1 no.1
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• pp.19-23
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• 2000

A novel recombinant baculovirus Autographa californica nuclear polyhedrosis virus (ACNPV) producing the green fluorescent polyhedra was constructed and characterized. The recombinant virus was stably produced fluorescent polyhedra in the infected cells and the morphology of the polyhedra was nearly similar to that of wild-type AcNPV. For the production of the fluorescent polyhedral the green fluorescent protein (GFP) gene was introduced under the control of polyhedrin gene promoter of AcNPV by translational fusion in the front and back of intact polyhedrin gene. The recombinant baculovirus was named as CXEP, As expected, the 93 kDa fusion protein was expressed in the CXEP-infected cells. Interestingly, however, the cells infected with CXEP also showed a 33 kDa protein band as cells infected with wild-type AcNPV. The results of Southern blot analysis and plaque assay suggested that two types of baculoviruses expressing the GFP fusion protein or only native polyhedrin were formed through homologous recombination between two polyhedrin genes in the same orientation. Thus, this system can be applied for the production of recombinant polyhedra with foreign gene product of diverse interest.

• International Journal of Industrial Entomology and Biomaterials
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• v.3 no.1
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• pp.75-81
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• 2001

We have constructed a novel recombinant Bombyx mori nuclear polyhedrosis virus (BmNPV) producing the green fluorescent polyhedra. For the production of the fluorescent polyhedra, partial polyhedrin gene containing KRKK as nuclear localization site from the BmNPV polyhedrin gene and the green fluorescent protein (gfp) gene were introduced under the control of p10 promoter of BmNPV. The recombinant BmNPV was stably produced fluorescent polyhedra in the infected Bm5 cells and the morphology of the fluorescent polyhedra was similar to that of wild-type BmNPV. The fluorescent polyhedra had 32 kDa native polyhedrin and 41 kDa fusion protein. From these data, we have further developed a novel BmNPV p10-based transfer vector producing recombinant polyhedra with foreign gene Product. The novel BmNPV P10-based transfer vector is composed of partial polyhedrin gene, factor Xa, and multiple cloning sites.

• Journal of Industrial Technology
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• v.29 no.B
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• pp.115-119
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• 2009

Green fluorescent protein (GFPuv) was displayed on the surface of ethanol-producing bacteria Zymomonas mobilis using N-terminal domain of ice nucleation protein (INP) as an anchoring motif. To evaluate the ice nucleation protein as plausible anchor motif in Z. mobilis, GFPuv gene was subcloned into Zymomonas expression vector yielding pBBR1MCS-3/pPDC/INPN/GFPuv plasmid., INP-GFPuv fusion protein was expressed in Z. mobilis and its fluorescence was verified by confocal microscopy. The successful display of GFPuv on Zymomonas mobilis suggest that INP anchor motif could be used for future fusion partner in Z. mobilis strain improvement.

• Fisheries and Aquatic Sciences
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• v.17 no.4
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• pp.479-486
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• 2014

To evaluate their potential utility as an ornamental organism, novel transgenic marine medaka Oryzias dancena strains with a highly vivid fluorescent phenotype were established through transgenesis of a cyan fluorescent protein gene (cfp) driven by the endogenous fast skeletal myosin light chain 2 gene (mlc2f) promoter. The transgenic marine medaka strains possessed multiple copies of transgene integrants and passed their fluorescent transgenes successfully to subsequent generations. Transgenic expression in skeletal muscles at both the mRNA and phenotypic levels was, overall, dependent upon transgene copy numbers. In the external phenotype, an authentic fluorescent color was dominant in the skeletal muscles of the transgenic fish and clearly visible to the unaided eye. The phenotypic fluorescent color presented differentially in response to different light-irradiation sources; the transgenics displayed a yellow-green color under normal daylight or white room light conditions, a strong green-glowing fluorescence under ultraviolet light, and a cyan-like fluorescence under blue light from a light-emitting diode.

• Proceedings of the Korean Institute of Information and Commucation Sciences Conference
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• 2014.10a
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• pp.977-980
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• 2014

A baculovirus vector systems including genes of polyhedron promoter, vesicular stomatitis virus G (VSVG), polyA, cytomegalovirus (CMV) promoter, enhanced green fluorescent protein (EGFP), and protein transduction domain (PTD) were constructed. These recombinant baculovirus vector systems were transfected into human foreskin fibroblast cells and various tissues and investigated gene transfer and expression of these vector systems with control vectors. From the study, these recombinant baculovirus vector systems were more effective and safe than control vector in view of gene transfer and expression.

• Journal of Life Science
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• v.17 no.2 s.82
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• pp.293-297
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• 2007

Green fluorescent protein (GFP) is an attractive reporter for bioprocess monitoring. A fluorescence-based method was developed to quantify GFP levels in transgenic plants and protein extracts. In this study, GFP was produced and secreted from suspension cells derived from transgenic rice. The RAmy3E promoter placed before the GFP gene controlled by sugars such as sucrose. The effects of sucrose concentration on the secretion of GFP and total protein into the medium were investigated in batch suspension culture. It was possible, therefore, to induce the expression of the GFP by removing sucrose from the cultured media or by allowing the rice suspension cells to deplete sucrose catabolically. The dry cell weight (7.06 g/L) and GFP level were detected as highest at 12%, 3% sucrose after 20 day culture, respectively. However secreted GFP fluorescence at the other sucrose concentrations (6%, 12%, 18% and 24%) were a little amount in media.

• Biotechnology and Bioprocess Engineering:BBE
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• v.7 no.5
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• pp.311-315
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• 2002

By the in vitro combinatorial mutagenesis, which is a sequential reaction of PCR mutagenesis and in vitro coupled transcription/translation with Escherichia coli S30 extract, S65 and E222 of green fluorescent protein of Aequarea victoria were comprehensively changed to all possible combinations of amino acids, thus totally 400 mutant (including a wild type) proteins were simultaneously produced and their fluorescent properties were analyzed. Although a few mutations had been reported so far at the 222nd position, replacement E222 to all other19 amino acids gave fluorescent signal to the mutants by changing Ser 65 to Ala together. Among the mutants, replacement to G, A, S, Q, H and C gave relatively high fluorescence. The in vitro combinatorial mutagenesis, therefore, has been proved valuable for comprehensive structure-function studies of proteins.

• Proceedings of the Korean Institute of Information and Commucation Sciences Conference
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• 2013.05a
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• pp.1006-1008
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• 2013

A novel recombinant baculovirus vector system containing coding genes for polyhedron promoter, vesicular stomatitis virus G (VSVG), polyA, cytomegalovirus (CMV) promoter, enhanced green fluorescent protein (EGFP), and protein transduction domain (PTD) was constructed. We applied this recombinant baculovirus vector into cells and murine tissues and compared efficacy of gene transfer and expression of this recombinant baculovirus vector system with control vector system. From this result, we confirmed that this novel recombinant baculovirus vector system was very effective than control vector system.