• Korean Journal of Microbiology
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• v.28 no.2
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• pp.91-97
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• 1990

Complementary DNA (cDNA) coding for human cytoplasmic superoxide dismutase (SOD1) (superoxide: superoxide oxidoreductase E.C.1.15.1.1) was isolated from human liver cDNA library of $\lambda$gt11 by in situ plaque hybridization. The insery cDNA gas the 5' untranslational region (UTR) and 3'UTR of SOD1 gene. Polymerase Chain Reaction (PCR) method was used fro subcloning of SOD1 structural gene. Using synthetic sense strand primer (24mer) containing a start codon and antisense strand primer (24mer), SOD1 structural gene was selectively amplified. Amplified DNA was directly cloned into the HincII site of pUC19 plasmid. Insery cDNA was subcloned into M13 mp19 and sequenced by dideowy chain termination method with Sequenase. The nucleotide sequence of insert cDNA had an open reading frame (ORF) coding for 153 amino acid residues. The structural gene of cytoplasmic SOD was placed under the control of bacteriophage $\lambda P_{L}$ regulatory sequences, generating a highly efficient expression plasmid. The production of human SOD1 in E. coli cells was about 7% of total cellular proteins and recombinant human SOD1 possessed its own enzymatic acitivity.

• Korean Journal of Microbiology
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• v.54 no.2
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• pp.126-135
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• 2018

The characteristics of enzyme and gene for mannanase B had been reported from Cellulosimicrobium sp. YB-43 producing some kind of mannanase. A gene coding for the enzyme, named mannanase C (ManC), was expected to be located downstream of the manB gene. The manC gene was cloned by polymerase chain reaction and sequenced completely. From this nucleotide sequence, ManC was identified to consist of 448 amino residues and contain a carbohydrate binding domain CBM2 besides a catalytic domain, which was homologous to mannanase belonging to the glycosyl hydrolase family 5. The catalytic domain of ManC showed the highest amino acid sequence similarity of 55% with the mannanases from Streptomyces sp. SirexAA-E (55.8%; 4FK9_A) and S. thermoluteus (57.6%; BAM62868). The His-tagged ManC (HtManC) lacking N-terminal signal peptide with hexahistidine at C-terminus was produced and purified from cell extract of recombinant Escherichia coli. The purified HtManC showed maximal activity at $65^{\circ}C$ and pH 7.5, with no significant change in its activity at pH range from 7.5 to 10. HtManC showed more active on konjac and locust bean gum (LBG) than guar gum and ivory nut mannan (ivory nut). Vmax and Km values of the HtManC for LBG were 68 U/mg and 0.45 mg/ml on the optimal condition, respectively. Mannobiose and mannotriose were observed on TLC as major products resulting from the HtManC hydrolysis of mannooligosacharides. In addition, mannobiose and mannose were commonly detected as the hydrolyzed products of LBG, konjac and ivory nut.

• Korean journal of applied entomology
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• v.38 no.2
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• pp.145-149
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• 1999

To develop the baculovirus expression vector system (BEVS) adopting p10 gene promoter of Spodoptera exigua nucleopolyhedrovirus (SeNPV), we characterized the p10 gene of SeNPV. The nucleotide sequence of 545 bases including the coding region of p10 gene was determined. Compared with the previously reported SeNPV p10 gene (Zuidema et al., 1993), 4 bases were different in the 5' and 3' flanking region but no difference was found in the coding region. The p10 gene was located within Xho I 1.5 Kb, Sph 1 2.4 Kb and Cla I 4.0 Kb fragments by Southern hybridization analysis. Also, the Sph I 2.4 Kb and the Cla I 4.0 Kb fragments were cloned and their restriction enzyme maps were determined.

• Journal of Life Science
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• v.14 no.4
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• pp.650-656
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• 2004

Oryza grandiglumis (CCDD, 2n=48), one of the wild rice species, has been known to possess fungal-,bacterial-, and insect-resistance against sheath blight, rice blast, bacterial leaf blight and brown plant hopper (Nilaparvata lugens). To rapidly isolate differentially expressed genes responding to fungal and wounding stress, wounding and yeast extract were treated to O. grandiglumis for 24 hrs. Suppression subtractive hybridization (SSH) method was used to obtain differentially expressed genes from yeast extract and wounding treated plants. Seven hundreds and seventy six clones were obtained by subcloning PCR product, and colony array and screening were carried out using radio-isotope labeled cDNA probes prepared from the wounding and yeast extract treated plants. One hundred and fifteen colonies were confirmed as true positive ones. Average insert size of the clones were ranged from 400 bp to 700 bp and all the inserts were sequenced. To decide the identity of those clones, sequences were analyzed by sequence homology via GenBank database. The homology search result showed that 68 clones were matched to the genes with known function; 16 were related to primary metabolism, 5 to plant retrotransposons, 5 to defense related metallothionein-like genes. In addition to that, others were matched to various genes with known function in amino acid synthesis and processing, membrane transport, and signal transduction, so on. In northern blot analysis, induced expressions of ogwfi-161, ogwfi-646, ogwfi-663, and ogwfi-695 by wounding and yeast extract treatments were confirmed. The result indicates that SSH method is very efficient for rapid screening of differentially expressed genes.

• Journal of Ginseng Research
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• v.14 no.2
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• pp.207-209
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• 1990

The sequence of ginseng peptide gene was designed and synthesized by the solid phase phosphoramidiate method. Synthetic segments were isolated, pllrified and joined to the plasmid pUC19. E.icherichiu coli JM101 cells were transformed with above hybrid plasmids. AmpiciIBin resistant transformants were screened and identified by in situ colony hybridization and Southern blot techniques. Finally the gene sequencing was done by the Sanger dideoxy method using primer extension.

• Proceedings of the Korean Aquaculture Society Conference
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• 2004.05a
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• pp.501-502
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• 2004

• Proceedings of the Korean Society of Applied Pharmacology
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• 1992.05a
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• pp.27-27
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• 1992

많은 홀몬, 성장인자 및 신경 전달물질들은 각각에 대한 세포막의 수용체와 결합하여 Phospholipase C (PLC)를 활성화시키므로서 신호를 세포내로 전달하여 세포의 성장, 대사, 신경 흥분, 수축 및 분비 등의 생리 현상을 나타내고 있다. 이 신호 전달의 중심 효소인 PLC는 현재까지의 효소 분리, 유전자 클로닝 등의 방법으로 3가지 Class에 적어도 8종유의 등위효소(Isozyme)들이 존재하고 있는 것으로 밝혀지고 있다. 본 연구에서는 이와 같은 등위효소에 특이적인 조절 물질을 선별할 수 있는 체계를 확립하기 위하여 새로운 동위효소의 분리 및 규명과 이 동위효소들에 대한 과발현 및 발현 세포주 개발을 추진하고 있다.

• Proceedings of the Korean Society of Crop Science Conference
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• 2001.05a
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• pp.66-67
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• 2001

• Proceedings of the Korean Society of Crop Science Conference
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• 1996.10a
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• pp.38-39
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• 1996

• Journal of Sericultural and Entomological Science
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• v.37 no.2
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• pp.181-185
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• 1995

To enhance expression of foreign gene by the novel expression vector, pBmKSK1, of Bombyx mori nuclear polyhedrosis virus, E. coli $\beta$-galactosidase gene expressing recombinant virus was infected in BmN-4 cells and various concentrations of silkworm hemolymph were added to the recombinant virus-infected BmN-4 cells containing fetal bovine serum. The expression efficiency of foreign gene was determined by $\beta$-galactosidase activity in the culture media. The results showed that the silkworm hemolymph was effective to expression of foreign gene in the BmN-4 cells, suggesting that the silkworm hemolymph could be substituted for fetal bovine serum in the BmN-4 cells to enhance expression of foreign gene.