• International Journal of Industrial Entomology and Biomaterials
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• v.6 no.1
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• pp.49-55
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• 2003

We describe the generation of transgenic silkworm that carrying the chimeric fibroin light chain (L-chain) gene. Previously, we have cloned the complete fibroin L-chain gene from the silkworm Baekok-Jam, Bombyx mori, and the complete fibroin gene from the oak silkworm, Antheraea yamamai. The 444 bp repetitive sequence of A. yamamai fibroin gene was inserted into the exon 6 of B. mori fibroin L-chain gene to produce chimeric fibroin L-chain gene. The chimeric fibroin L-chain gene was cloned into the polyhedrin gene site of Autographa californica nuclear polyhedrosis virus (AcNPV) to yield a recombinant baculovirus as a fibroin gene targeting vector, One-day-old fifth instar female silkworm larvae were injected with the recombinant baculovirus and then mated with normal male moths. Genomic DNA from their progenies was extracted and screened for the desired targeting event by using PCR and Southern blot analysis. The analysis showed that the chimeric fibroin gene had intergrated into the L-chain gene on the genome by homologous recombination and was transmitted through generations. The transgenic silkworm carrying the chimeric fibroin gene were approximately 43.2% in $F_2$ generation, and the silkworms synthesized the fusion protein in cocoons layer.

• Asian-Australasian Journal of Animal Sciences
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• v.23 no.7
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• pp.855-862
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• 2010

Among the known interferon-induced antiviral mechanisms, the Mx pathway is one of the most powerful pathways. The Mx protein has direct antiviral activity and inhibits a wide range of viruses by blocking an early stage of the viral replication cycle. Cloning, characterization, and expression of Mx in vivo and in vitro have been conducted. The chicken Mx gene spans 21 kb and is made up of 14 exons and 13 introns, of which the promoter region was analyzed. The real-time PCR results showed that Mx expression was increased in chicken embryo fibroblasts (CEF) after 12- and 24-h induction with polyI: C. Induction of Mx expression by poly I: C in vivo revealed tissue-specific patterns among the chicken tissues tested. A trace expression of Mx was detected in healthy chicken liver tissues from adult chickens without inducement; the expression levels in the liver, heart, and gizzard were higher than in the muscle and kidney. This is the first report to demonstrate the expression of a glutathione-S-transferase-tagged-Mx fusion protein of 75 KDa, as well as the biological activity tested by SDS-PAGE and western blotting.

• Journal of Microbiology and Biotechnology
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• v.16 no.6
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• pp.952-960
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• 2006

As a unicellular green alga that possesses many of the metabolic pathways present in higher plants, Chlorelia offers many advantages for expression of heterologous proteins. Since strong and constitutive promoters are necessary for efficient expression in heterologous expression systems, the development of such promoters for use in the Chlorella system was the aim of this study. Proteins encoded by the early genes of algal viruses are expressed before viral replication, probably by the host transcriptional machinery, and the promoters of these genes might be useful for heterologous expression in Chlorella. In this study, putative promoter regions of DNA polymerase, ATP-dependent DNA ligase, and chitinase genes were amplified from eight Korean Chlorella virus isolates by using primer sets designed based on the sequence of the genome of PBCV-1, the prototype of the Phycodnaviridae. These putative promoter regions were found to contain several cis-acting elements for transcription factors, including the TATA, CAAT, NTBBF1, GATA, and CCAAT boxes. The amplified promoter regions were placed into Chlorella transformation vectors containing a green fluorescence protein (GFP) reporter gene and the Sh ble gene for phleomycin resistance. C. vulgaris protoplasts were transformed and then selected with phleomycin. The GFP fluorescence intensities of cells transformed with chitinase, DNA polymerase, and DNA ligase gene promoter-GFP fusion constructs were 101.5, 100.8, and 95.8%, respectively, of that of CaMV 35S-GFP-transformed Chlorella cells. These results demonstrate that these viral promoters are active in transformed Chlorella.

• Journal of Plant Biotechnology
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• v.1 no.1
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• pp.46-55
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• 1999

The expressed sequence tags(ESTs) from immature seed of rice, Oryza sativa cv Milyang 23, were partially sequenced and analyzed by homology. As of 1998, the partial sequences of about 6,600 cDNA clones were analyzed from normal and normalized immature seed cDNA libraries. About 2,200 ESTs were putatively identified by BLASTX deduced amino acid sequence homology analysis. About 20% of them were putatively identified as storage proteins. Also the clones were highly homologous to genes involved particularly in starch biosynthesis, glycolysis, signal transduction and defenses. Compared to 35% of redundancy in the ESTs of normal cDNA library, that from the substracted library was 15%. The Korea Rice Genome Network is maintained to provide the updated information of sequences, their homologies and sequence alignments of ESTs. For the stable expression of transgene in rice, diverse vectors were developed for overexpression, targeting and gene dosage effect with transit peptides (Tp) and matrix attachment region (MAR) sequence from chicken lysozyme locus. The rice calli were transformed via Agrobacterium tumefaciens LBA4404(pSB1) with the triparental mating technique and selected by herbicide resistance. The green fluorescent protein(GFP) gene in expression vector under the control of rbcS promoter-Tp was overexpressed upto 10 % of the total soluble protein. In addition, the Tp-sGFP fusion protein was properly processed during translocation into chloroplast. The expression of sGFP in the presence of MAR sequences was analyzed with Northern and immunoblot analysis. All the lines in which sGFP transgene with MAR sequence, showed position independent and copy number-dependent expression, while the lines without MAR showed the varied level of expression with the integration site. Thus the MAR sequence significantly reduced the variation in transgene expression between independent transformants.

• Journal of Life Science
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• v.7 no.1
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• pp.49-58
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• 1997

Single P[en-lacZ] element including 5.7 kb of engrailed upstream sequences and the E. coli lacZ fusion gene, localized on 48A in rxyho25 strain was transposed to different sites in the Drosophila genome by the jumpstart technique. From 3315 individual genetic crosses, 113 new insertion lines carrying P[en-lacZ] inserted at different sites were obtained. $\beta$-Galactosidase expression in larval tissues of 113 insertion lines were detected by X-gal staining. & among 113 lines have been indentified to be for recessive lethal mutations. Among 7 lines, the #1119 line being lethal during embryogenesis was examined about the ${\beta}$$-Galactosidase expression, nuclear behavior and cellularization pattern during embryogenesis. The P[en-lacZ] insertion lines obtained in this study could be utilized for studying structure and function of the Drosophila development-related genes.

• Journal of Microbiology and Biotechnology
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• v.23 no.1
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• pp.56-63
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• 2013

We have characterized the putative ${\alpha}$-glucosidase gene (st2525) selected by total genome analysis from the acidothermophilic crenarchaeon Sulfolobus tokodaii strain 7. The ORF was cloned and expressed as a fusion protein in Escherichia coli, and recombinant ST2525 was purified by Ni-NTA affinity chromatography. Maximum activity was observed at $95^{\circ}C$ and pH 4.0, and the enzyme exhibited stability with half-lives of 40.1 min and 7.75 min at extremely high temperatures of $100^{\circ}C$ and $105^{\circ}C$, respectively. The enzyme retained at least 85% of its maximal activity in the pH range of 4.0-11.0. ST2525 exclusively hydrolyzed ${\alpha}$-1,4-glycosidic linkages of oligosaccharides in an exo-type manner, with highest catalytic efficiency toward maltotriose. The enzyme also displayed transglycosylation activity, converting maltose to isomaltose, panose, maltotriose, isomaltotriose, etc. From these results, ST2525 could be potentially useful for starch hydrolysis as well as novel synthesis of oligosaccharides in industry.

• Molecules and Cells
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• v.23 no.3
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• pp.316-322
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• 2007

Zinc finger transcription factor genes are a significant fraction of the genes in the vertebrate genome. Here we report the isolation and characterization of a human zinc finger-containing gene, ZNF435, from a fetal brain cDNA library. ZNF435 cDNA is 1290 base pairs in length and contains an open reading frame encoding 349 amino acids with four C2H2-type zinc fingers at its carboxyl terminus and a SCAN motif at its amino terminus. RT-PCR results showed that ZNF435 was expressed in all tested tissues. A ZNF435-GFP fusion protein was located in the nucleus and the four zinc fingers acted as nuclear localization signals (NLSs). ZNF435 was found to be capable of homo-association, and this effect was independent of its zinc fingers. Furthermore, ZNF435 proved to be a transcription repressor as its overexpression in AD293 cells inhibited the transcriptional activities of AP-1.

• Journal of Microbiology and Biotechnology
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• v.32 no.6
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• pp.816-823
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• 2022

The rapid spread of superbugs leads to the escalation of infectious diseases, which threatens public health. Endolysins derived from bacteriophages are spotlighted as promising alternative antibiotics against multi-drug resistant bacteria. In this study, we isolated and characterized the novel Salmonella typhimurium phage PBST08. Bioinformatics analysis of the PBST08 genome revealed putative endolysin ST01 with a lysozyme-like domain. Since the lytic activity of the purified ST01 was minor, probably owing to the outer membrane, which blocks accessibility to peptidoglycan, antimicrobial peptide cecropin A (CecA) was fused to the N-terminus of ST01 to disrupt the outer membrane. The resulting CecA::ST01 has been shown to have increased bactericidal activity against gram-negative pathogens including Pseudomonas aeruginosa, Klebsiella pneumoniae, Acinetobacter baumannii, Escherichia coli, and Enterobacter cloacae and the most affected target was A. baumannii. In the presence of 0.25 µM CecA::ST01, A. baumannii ATCC 17978 strain was completely killed and CCARM 12026 strain was wiped out by 0.5 µM CecA::ST01, which is a clinical isolate of A. baumannii and resistant to multiple drugs including carbapenem. Moreover, the larvae of Galleria mellonella could be rescued up to 58% or 49% by the administration of CecA::ST01 upon infection by A. baumannii 17978 or CCARM 12026 strain. Finally, the antibacterial activity of CecA::ST01 was verified using 31 strains of five gram-negative pathogens by evaluation of minimal inhibitory concentration. Thus, the results indicate that a fusion of antimicrobial peptide to endolysin can enhance antibacterial activity and the spectrum of endolysin where multi-drug resistant gram-negative pathogens can be efficiently controlled.

• IMMUNE NETWORK
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• v.21 no.4
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• pp.30.1-30.12
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• 2021

High expression of mitofusin-2 (MFN2), a mitochondrial fusion protein, has been frequently associated with poor prognosis of patients with cervical cancer. Here, we aimed to identify the function of MFN2 in cervical cancer to understand its influence on disease prognosis. To this end, from cervical adenocarcinoma, we performed an MTT assay and quantitative RT-PCR (qRT-PCR) analysis to assess the effects of MFN2 on the proliferation and of HeLa cells. Then, colony-formation ability and tumorigenesis were evaluated using a tumor xenograft mouse model. The migration ability related to MFN2 was also measured using a wound healing assay. Consequently, epithelial-mesenchymal transition (EMT) of MFN2-knockdowned HeLa cells originating from adenocarcinoma. markers related to MFN2 were assessed by qRT-PCR. Clinical data were analyzed using cBioPortal and The Cancer Genome Atlas. We found that MFN2 knockdown reduced the proliferation, colony formation ability, migration, and in vivo tumorigenesis of HeLa cells. Primarily, migration of MFN2-knockdowned HeLa cells decreased through the suppression of EMT. Thus, we concluded that MFN2 facilitates cancer progression and in vivo tumorigenesis in HeLa cells. These findings suggest that MFN2 could be a novel target to regulate the EMT program and tumorigenic potential in HeLa cells and might serve as a therapeutic target for cervical cancer. Taken together, this study is expected to contribute to the treatment of patients with cervical cancer.

• Journal of Mushroom
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• v.14 no.4
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• pp.142-154
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• 2016

Worldwide production of mushrooms has been increasing by 10-20% every year. Recently, Pleurotus eryngii and P. nebrodensis have become popular mushroom species for cultivation. In particular, China exceeded 8.7 million tons in 2002, which accounted for 71.5% of total world output. A similar trend was also observed in Korea. Two kinds of mushrooms-Gumji (金芝; Ganoderma) and Seoji-are described in the ancient book 'Samguksagi' (History of the three kingdoms; B.C 57~A.D 668; written by Bu Sik Kim in 1145) during the Korea-dynasty. Many kinds of mushrooms are also described in more than 17 ancient books during the Chosun-dynasty (1392~1910) in Korea. Approximately 200 commercial strains of 38 species of mushrooms were developed and distributed to cultivators. The somatic hybrid variety of oyster mushroom, 'Wonhyeong-neutari,' was developed by protoplast fusion, and distributed to growers in 1989. Further, the production of mushrooms as food was 199,829 metric tons, valued at 850 billion Korean Won (one trillion won if mushroom factory products are included) in 2015. In Korea, the major cultivated species are P. ostreatus, P. eryngii, Flammulina velutipes, Lentinula edodes, Agaricus bisporus, and Ganoderma lucidum, which account for 90% of the total production. Since mushroom export was initiated in 1960, the export and import of mushrooms have increased in Korea. Technology was developed for liquid spawn production, and automatic cultivation systems led to the reduction of production cost, resulting in the increase in mushroom export. However, some species were imported owing to high production costs for effective cultivation methods. In academia, RDA scientists have conducted mushroom genome projects since 1997. One of the main outcomes is the whole genome sequencing of Flammulina velutipes for molecular breeding. With regard to medicinal mushrooms, we have been conducting genome research on Cordyceps and its related species for developing functional foods. There are various kinds of beneficial substances in mushrooms; mushroom products, including pharmaceuticals, tonics, healthy beverages, functional biotransformants, and processed foods have also became available on the market. In addition, compost and feed can likewise be made from mushroom substrates after harvest.