• Korean Journal of Veterinary Research
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• v.37 no.4
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• pp.875-882
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• 1997

To produce the recombinant fibronectin-binding protein(FnBP) for development of subunit vaccine against Staphylococcus aureus. The fnbp gene was amplified from the chromosomal DNA of S aureus KNU 196 strain using the polymerase chain reaction, and cloned into pGEX-4T-2. Then, the recombinant FnBP fused with glutathione-S-transferase was produced in E coli, purified by affinity chromatography, and identified its antigenicity and immunogenicity by Western blot. The recombinant FnBP produced in this study is considered to have the same property of native FnBP purified from S aureus, and is expected to be useful as a candidate for S aureus subunit vaccine.

• Journal of fish pathology
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• v.26 no.3
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• pp.231-240
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• 2013

Adjuvants are immune enhancers that are often used in vaccination to augment the immune response of a vaccine, thereby enhancing the protective immunity against the targeted disease. In the present study, we used the recombinant protein, such as rRbCC1, this protein was produced from rock bream CC chemokine 1. To verify the adjuvant effects of this recombinant protein, the immune responses of rock bream to Streptococcus iniae (S. iniae) FKC vaccination, which alone or in combination with recombinant protein was analyzed and then also performed experimental challenge with live S. iniae. The result of serum agglutination titres was showed relatively low levels however, the efficacy of FKC vaccine still conferred protection against S. iniae. Moreover, the adverse effects result showed that no statistically significant difference was revealed between high concentration injected and non-injected fish groups, generally. The relative percent survival (RPS) of FKC + recombinant vaccination group was significantly higher than that of vaccinated group with FKC alone. After experimental challenge to the rock bream by injection with live bacteria (S. iniae), the FKC + rRbCC1 vaccination group was showed 87.0% RPS, however, the RPS of FKC alone vaccination was 68.2%. The results indicated that the recombinant protein as an adjuvant had a clear synergism to injection vaccine of rock bream.

• Journal of Microbiology and Biotechnology
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• v.23 no.8
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• pp.1116-1122
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• 2013

Cell line development is the most critical and also the most time-consuming step in the production of recombinant therapeutic proteins. In this regard, a variety of vector and cell engineering strategies have been developed for generating high-producing mammalian cells; however, the cell line engineering approach seems to show various results on different recombinant protein producer cells. In order to improve the secretory capacity of a recombinant tissue plasminogen activator (t-PA)-producing Chinese hamster ovary (CHO) cell line, we developed cell line engineering approaches based on the ceramide transfer protein (CERT) and X-box binding protein 1 (XBP1) genes. For this purpose, CERT S132A, a mutant form of CERT that is resistant to phosphorylation, and XBP1s were overexpressed in a recombinant t-PA-producing CHO cell line. Overexpression of CERT S132A increased the specific productivity of t-PA-producing CHO cells up to 35%. In contrast, the heterologous expression of XBP1s did not affect the t-PA expression rate. Our results suggest that CERT-S132A-based secretion engineering could be an effective strategy for enhancing recombinant t-PA production in CHO cells.

• The Microorganisms and Industry
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• v.19 no.4
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• pp.14-26
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• 1993

Optimizing protein production in recombinant E. coli strains involves manipulation of genetic and environmental factors. In designing a production system, attention must be paid to gene expression efficiency, culture conditions and bioreactor configuration. Although not much emphasis was given to the physiology of host strains in this review, an understanding of the relationship between the physiology of host cell growth and the overproduction of a cloned gene protein is of primary importance to the improvement of the recombinant fermentation processes. Sometimes it is desirable to make use of gene fusion systems, e.g. protein A, polypeptide, gutathione-S-transferase, or pneumococcal murein hydrolase fusion, to facilitate protein purification.

• IMMUNE NETWORK
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• v.2 no.3
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• pp.175-181
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• 2002

Background: S100A6 is a calcium-binding protein overexpressed in several tumor cell lines including melanoma with high metastatic activity and involved in various cellular processes such as cell division and differentiation. To detect S100A6 protein in patient' samples (ex, blood or tissue), it is essential to produce a monoclonal antibody specific to the protein. Methods: First, cDNA coding for ORF region of human S100A6 gene was amplified and cloned into the expression vector for GST fusion protein. We have produced recombinant S100A6 protein and subsequently, monoclonal antibodies to the protein. The specificity of anti-S100A6 monoclonal antibody was confirmed using recombinant S100A recombinant proteins of other S100A family (GST-S100A1, GST-S100A2 and GST-S100A4) and the cell lysates of several human cell lines. Also, to identify the specific recognition site of the monoclonal antibody, we have performed the immunoblot analysis with serially deleted S100A6 recombinant proteins. Results: GST-S100A6 recombinant protein was induced and purified. And then S100A6 protein excluding GST protein was obtained and monoclonal antibody to the protein was produced. Monoclonal antibody (K02C12-1; patent number, 330311) has no cross-reaction to several other S100 family proteins. It appears that anti-S100A6 monoclonal antibody reacts with the region containing the amino acid sequence from 46 to 61 of S100A6 protein. Conclusion: These data suggest that anti-S100A6 monoclonal antibody produced can be very useful in development of diagnostic system for S100A6 protein.

• Journal of Microbiology and Biotechnology
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• v.10 no.1
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• pp.115-118
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• 2000

Recombinant plasmids harboring a heterologous gene coding for the enhanced green fluorescent protein (EGFP) were transfected and expressed in Drosophila melanogaster S2 cells. A stable transformation of polyclonal cell populations expressing EGFP were isolated after 4 weeks of selection with hygromycin B. The recombinant EFGP expressed in transformed S2 cells consisted of a molecular weight of 27 kDa. EGFP expression was also confirmed by fluorometric measurement. The maximum EGFP concentration was about 9.3 mg/I. The present findings demonstrate not only the successful stable expression of EGFP in Drosophuila was about 9.3 mgI. The present findings demonstrate not only the successful stable expression of EGFP in Drosophila S2 cells, but also the use of EGFP as a reporter to analyze gene expression, with its potential of a Drosophila cell expression system for recombinant protein production being an alternative to a baculovirus-insect cell expression system.

• Journal of Microbiology and Biotechnology
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• v.12 no.4
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• pp.563-568
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• 2002

Stably transformed Drosophila melanogaster 52 cells producing recombinant VP7 were obtained, and recombinant VP7 expression was confirmed by Western blot analysis. The molecular weight of recombinant VP7 expressed in 52 cells was approximately 35.5 kDa, and 75% of the total VP7 produced was present in the medium. Recombinant VP7 contained N-linked glycosylated oligosaccharides. Aprotinin, leupeptin, and polyvinylpyrrolidone did not have any noticeable effect on recombinant VP7 production; however, DMSO and sodium butyrate increased its production by 120% and 60%, respectively.

• IMMUNE NETWORK
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• v.1 no.3
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• pp.187-195
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• 2001

The expression of recombinant proteins fused to 26 kDa glutathione S-transferase (GST) extracted from Schistosoma japonicum represents an attractive system for purifiying proteins of interest in a single step using GST-affinity chromatography. In addition, the GST-tag is used conveniently for detecting fused proteins since its high solubility as well as its relatively small size rarely interferes with the biological activity of the fused protein. In this regard, the GST system is frequently applied for tracing fusion proteins in both prokaryotic and eukaryotic cells to elucidate the physiological interactions and functional compartments of proteins. To provide a further tool in analyzing GST-fusion proteins, a new monoclonal antibody, with a high specificity to the S. japonicum GST was produced. Methods: BALB/c mice were immunized both with recombinant S. japonicum GST proteins, and by the fusion of splenocytes from these mice with myeloma cells. From this, a new anti -GST monoclonal antibody, termed SARAH, was generated. The specificity and reactivity of this antibody was confirmed by ELISA and by Western blot analysis. Results: SARAH showed a high reactivity to recombinant GST and GST fusion protein but not with native mammalian GST proteins as derived from other species including humans, cows, rabbits and rats. The applicability of SARAH was further demonstrated by confocal laser scanning microscopy, where GST proteins that were expressed transiently in mouse fibroblast cells, were specifically detected without interference of endogenous GST. Conclusion: SARAH is new monoclonal antibody with a high specificity to recombinant GST proteins but not to endogenous GST in mammalian cells.

• KSBB Journal
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• v.18 no.6
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• pp.435-439
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• 2003

Plant cell culture can be divide into two classes non-organic culture and organic culture. Non-organic culture such as suspension culture has many researches, however organic culture about recombinant protein production has little researches. Recombinant protein produced through organ culture is quite stable and it can make proteins by itself without any grow regulators. Therefore organ culture is much easier than other methods. In this research, we used transformed tobacco seed. At first we germinated the seed then separated stems and leaves from the grown plant. And raised in liquid medium by in vitro vegetative reproduction. Continuing most suitable conditions, we compared the Quantities of recombinant protein from intra cellular with from extra cellular. And adding some permeabilizing agents (Pluronic F-68, Triton X-100, DMSO, PEG8000), we increased the productivity of the recombinant protein.

• KSBB Journal
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• v.11 no.4
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• pp.438-444
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• 1996

Using recombinant Vaccinia virus(vSC8) that express ${\beta}$-galactosidase, a model heterologous protein, conditions for virus and protein production were investigated in tissue culture flask. As host animal cells HeLa and HeLa S3 were used. It was demonstrated that cells infected during the exponential growth phase gave higher protein yield than those infected during the stationary growth phase and calf serum concentration after virus infection did not significantly alter protein yield. Pretreatment of cell layer with hypotonic solution enhanced the virus infectivity. Optimum cell growth and recombinant protein production was achieved at $37^{\circ}C$. But, during 2 hours of virus infection period incubation temperature must be lowered to 20∼$30^{\circ}C$ for maximum recombinant protein yield. To enhance virus replication, the effects of adrenal glucocorticoid hormone (Dexamethasone) and silkworm hemolymph were evaluated. Only dexamethasone increased about 20% of ${\beta}$-galactosidase yield in HeLa S3 cells when added with 10-7∼10-5M concentration 24 hours before infection.