• Journal of Fisheries and Marine Sciences Education
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• v.26 no.4
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• pp.808-822
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• 2014

넙치 (Paralichthys olivaceus)로부터 elastase-like serine protease (PoElSp)를 암호화하는 cDNA를 클로닝하여 그 서열을 분석한 결과, PoElSp 유전자는 269 아미노산을 암호화하는 978염기쌍으로 구성되었다. PoElSp 유전자의 조직 특이적 발현 양상을 RT-PCR법으로 조사한 결과, 간, 비장 및 소장에서 그 발현이 크게 나타났다. lipopolysaccharide (LPS)로 인위적 세균감염을 유도한 후, 1시간째에 콩팥에서, 3시간째에는 근육에서, PoElSp 유전자의 발현이 크게 증가하였다. 또한, 이 유전자의 발현은 비장에서 LPS 주입 후 1-24시간동안 점차로 증가하였다. pro-mature PoElSp (proPoElSp)에 해당하는 cDNA를 pET32a 벡터 시스템을 이용하여 대장균에서 발현시켰다. 이 재조합 proPoElSp 단백질의 활성은 gelatin zymography 방법과 합성형광 Z-Phe-Arg-AMC의 분해법을 이용하여 측정하였다. 단백질 분해효소 활성을 위한 최적 pH는 7.5였다. 실험결과들을 종합하면, PoElSp 단백질은 면역 반응에서 중추적 역할을 하리라 판단된다.

• Korean Journal of Microbiology
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• v.30 no.4
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• pp.246-251
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• 1992

Molccular cloning of nod genes from Bradvrhizobium sp. SNU001, a nitrogen-fixing symbiont isolated from thc root nodules of soybean (Clycine trim) . was carried out. nod genes were found to be located on thc genome of the symbiont by gcnomic hybridization with 4.5 kb EcoRI/HndIII fragment (nod DABC) of Rhizohium meliloti as probe. Genomic library of this symbiont was constructed using h phage EMBL3-BanlHI vector. from which five nod positive clones were sclectcd by primary and secondary screening methods. The partial restriction map of inserted genomic DNA of h CNS-l(c1one 2) was constructed. and 3.9 kh Bun7HI fragment. which showed strong hybridization signal to the probe, was subcloned into pBS KS(+) plasmid vector. Partial restriction inap ot' a selected subclone (pBjCNS-I) was constructed and nod DABC was found to be located on the 1.8 kb KpnI/Sacl fragment of this subclone.

• Korean Journal of Microbiology
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• v.35 no.3
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• pp.226-230
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• 1999

To characterize the Korean isolate of infeciious hematopoietic necrosis virus (IHNV-PRT), a partial DNA fragment G gene of the MNV-PRT was amplified by RT-PCR. cloned inlo pGEM-T easy vector and analyzed for nucleotlde sequences. The size of the PCR pmduct was about 442 bp. The nucleotlde sequence homologies ofthe G gene of IHNV-PRT were 95%, 94%, 94% 94%, 93%, 53%. respectively. with those of foreign isolates of IHNV, IHNV-RB-76. IHNV-LR-73, MNV-K, IHNV-WRAC, Im-SRCV, IHNV-Col-85. However, it showed 81% homology with that of other fish rhabdovirus, hisame rhabdovirus (HRV). Frou~ the rcsults of deduced amino acid sequence homology analysis. G protein of IHNV-PRT showed 96% hornologies with those of foreign isolates of IHNV but 89% homology with that of HRV These results indicaled that, even though G gene of IHIW-PRT showed low homology with that of HRY it was highly conserved among different strains of THNV.

• Journal of the Korea Institute of Information and Communication Engineering
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• v.17 no.4
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• pp.1005-1009
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• 2013

Using DNA sequencing method, we analyzed mutations of rpoB (RNA polymerase beta subunit) rifampin-resistant Mycobaterium tuberculosis strains which were identified by conventional test at Masan National Hospital and The Korean Institute of Tuberculosis. Though it has been reported different mutations of rpoB region of rifampin-resistant M. tuberculosis strains in the south of Korea, it is not confirmed whether these mutations of rpoB region actually express rifampin resistance through experiment. We confirmed experimentally these mutations of rpoB region of M. tuberculosis strains induced rifampin-resistance through ampified rpoB by polymerase chain reaction (PCR) and cloning of mutant rpoB into rifampin sensitive-M. tuberculosis strain.

• Korean Journal of Microbiology
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• v.46 no.2
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• pp.207-212
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• 2010

A bacterium producing the extracellular mannanase was isolated from Korean soybean paste. The isolate WL-8 has been identified as Bacillus subtilis on the basis on its 16S rRNA sequence, morphology and biochemical properties. The mannanase productivity of strain WL-8 was increased in LB broth by addition of wheat bran. The maximum mannanase productivity was reached to approximately 20 U/ml in LB medium supplemented with 6% wheat bran. A gene encoding the mannanase of WL-8 was cloned into Escherichia coli and its nucleotide sequence was subsequently determined. The mannanase gene consisted of 1,086 nucleotides encoding a polypeptide of 362 amino acid residues. The deduced amino acid sequence was highly homologous with those of several mannanases from B. subtilis belonging to GH family 26. Reaction temperature and pH profiles were investigated using the culture filtrate and cell-free extract of the recombinant E. coli carrying a WL-8 mannanase gene, respectively. Optimal conditions for the two fractions occurred at pH 5.5 and $60^{\circ}C$. The cell-free extract showed higher mannanase activity than the culture filtrate at above $60^{\circ}C$.

• The Journal of Natural Sciences
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• v.5 no.1
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• pp.37-45
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• 1992

The gene coding for $\beta$-galactosidase of Neisseria lactamica 2118 was cloned into Escherichia coli MC 1061. The isolated 6.5 Kb EcoR I fragement and 7.2 Kb BamH I fragment of chromosomal DNA in Southern hybridization were ligated to a vector plasmid pBR322 and then transformed into Escherichia coli MC 1061 cells. Finally, I obtained three clones as $\beta$-galactosidase positive clone by colony hybridization and Southern hybridization($\beta$-galactosidase probe: lac Z gene of pMC1871). Three recombinant plasmids(pNL.13. 17 and 24) were found to contain the 7.2Kb BamH I fragment originated from Neisseria lactamica 2118 chromosomal DNA by Southern hybridization and pNL 24 was showed high homology to probe especially and also its physical map was constructed.

• Microbiology and Biotechnology Letters
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• v.20 no.3
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• pp.280-288
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• 1992

Molecular cloning of metalloprotease gene from Serratia marcescens ATCC 21074 into Escherichia coli JM109 was carried out. Chromosomal DNA of S. marcescens was completely digested with Hind111 and southern hybridization with a synthetic oligonucleotide probe revealed that a 50 KD metalloprotease gene was contained in 4.0 Kb chromosomal DNA fragment, 4.0 Kb chromosomal DNA fragments eluted from agarose gel were ligated with pUC19 and transformed into E. coli JM109. Nine positive clones were obtained from about $1\times 10^3$ transformants by colony hybridization. Their recombinant plasmids, pSPl and pSP2 have same chromosomal DNA fragments in pUC19 in opposite-orientations. When cloned metalloprotease gene was expressed in E. coli, about 52 KD precursor protein of metalloprotease was detected by western blot analysis from E. coli harboring a recombinant plasmid pSP2. Plasmid pSP2 showed no protease activities in E. coli but overproduced the active metalloprotease in S. rnarcescens ATCC 27117.

• Korean Journal of Microbiology
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• v.32 no.1
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• pp.28-33
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• 1994

In normal cells tau protein is associated with axonal microtubules, whereas in Alxheimer's disease it is immobilized in the somatodendritic compartment of certain nerve cells as a major component of the paired helical filament. As a part of the study to analyze the nature of the paired helical filament (PHF) deposits and some related factors in brain, we have cloned and expressed a human tau gene cDNA in Escherichia coli to obtain the recombinant human tau protein in abundance.

• Korean Journal of Microbiology
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• v.30 no.6
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• pp.539-545
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• 1992

Recombinant plasmid pPLac15 determined both phosphoenolpyruvate-dependent phosphotransferase uptake of lactose and phospho-$\beta$-galactosidase (Moon et al., 1989). A restriction mapping of the pPLac15 was compiled with several restriction enzymes and a seriese of sub clones into pUC18 was constructed. From an analysis of the proteins produced by Escherichia coli cells of transformants containing each of the recombinant subclone plasmids, it was found that the gene for phospho-$\beta$-galactosidase in pUCI8 was expressed about 1.8-folds in E. coli.

• Microbiology and Biotechnology Letters
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• v.20 no.2
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• pp.129-135
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• 1992

A chitinase gene of Serratia marcescens ATCC 27117 was cloned and expressed in Escherichiu di. A genomic library of S, marcescens was constructed with pUC 19 and screened using the swollen chitin agar plate for chitinolytic clones. A positive clone showing chitinclearance contains a recombinant pCHI 89, composed of 8.9 Kb chromosomal DNA fragment and pUC 19. Plasmid pCHI 89 produced 58 KD chitinase in E. coli, which was coincided with one of five extracellular chitinases produced by S. nzarccscens. Restriction endonuclease cleavage sites of the 8.9 Kb insert DNA fragment were mapped. E. coli JM109 harboring pCHI 89 inhibits the growth of a plant pathogenic fungus, Fusarium oxysporum.