• Microbiology and Biotechnology Letters
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• v.26 no.2
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• pp.110-116
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• 1998

The secY gene of Streptomyces lividans TK24 was cloned by the PCR method with synthetic oligonucleotide primers designed on the basis of the conserved regions of Ll5-secY-adk operon from E. coli, B. subtilis, and M luteus. The deduced amino acid sequences of the SecY are highly homologous to those of other known SecY. It has 46%, 43%, 57%, 44%, 42%,56%, 90% similarity to Escherichia coli, Bacillus subtilis, Micrococcus luteus, Bacillus licheniformis Staphylococcus carnosus, Brevibacterium flavum, Streptomyces scabies, respectively and almost the same with Streptomyces coelicolor, The gene organization of Ll5- SecY-Adk is also similar to those of other bacteria. SecY and Adk are very likely translationally coupled that is overlapping stop codon of SecY and start codon of Adk with one base pair, which is common structure among high GC content strains of gram positive bacteria.

• Journal of the Korea Organic Resources Recycling Association
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• v.11 no.3
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• pp.67-74
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• 2003

Starvation promoters can be utilized for in situ bioremediation and for the efficient bioprocessing. Previously we have cloned and characterized strong starvation promoters from envrionmentally relevant bacteria, Pseudomonas putida strains (Y. Kim, and A. Matin, J. Bacteriol. 177:1850-1859, 1995). Here we report the construction of the plasmid pYKS101 using one of the starvation promoters from P. putida MK1. The pYKS101 was found to be useful for a novel starvation promoter-driven gene expression system. Under this system, the target reporter gene, lacZ, was stably integrated into the chromosomal DNA of P. putida MK1. ${\beta}$-galactosidase activity was found to be four-fold higher upon carbon starvation than during exponential growth. The resultant recombinant strain is indigenous to the environment contaminated with various toxic materials, hence can be a good candidate for in situ bioremediation.

• Korean Journal of Microbiology
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• v.33 no.2
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• pp.92-96
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• 1997

The 6.8 kb Xhol fragment of chromosomal ONA of Pseudomonas sp. 0177 contains the phnDEFG genes involved in the degradation of polyaromatic hydrocarbons and chlorinated aromatics. Here, we report the nucleotide sequence of the ORF encoding a polypeptide consisted of 143 amino acids with a Mr of 13,859. The nucleotide sequence of the ORF is 99% and 68.6% identical to the downstream region of catE of Sphingomonas sp. strain HV3 and the ORF between xylE and xylG of Sphingomonas yanoikuyae Bl, respectively. The deduced amino acid sequence of the PhnF has 62.3% identity with the amino acid encoded hy orfY region of Citrobacter freundii DSM30040. We now confirm that the ORF is located between the catechol 2,3-dioxygenase (C230), phnE, and 2-hydroxymuconic semialdehyde dehydrogenase (2HMSO), phnG.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1994.04a
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• pp.192-192
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• 1994

본 논문에서는 cytochrome P450 LA1 유전자의 5'-upsteam 조절부위의 클로닝을 실시하였다. pUC19 vector에 연결시킨 3.4 Kb 크기의 Pstl DNA조각을 Sst1, Nco1 제한 효소로 자른 뒤, Exonuclease III 를 처리하여 약 200bp 씩의 차이를 갖는 여러 크기의 plasmid들을 얻었다. 이 plasmid 의 핵산서열을 알아보기 위해 dideoxy nucletide를 이용한 sequencing방법으로 그 핵산서열의 결정 실험을 시도하였다. 또한, 다환상 방향족 탄화수소 화합물에 반응성을 갖는 C57BL/6N 생쥐와 반응성을 갖지않는 DBA/2N 생쥐에 있어 phase II 대사 효소인UDP-glucuronosyltransferase 효소활성에 대한 3-methylcholanthrene의 영향을 알아보기 위해 C57BL/6N 생쥐와 DBA/2N 생쥐에 각각 다른 농도의 3-methylcholanthrene을 처리하거나 각기 다른 시간에 3-methylcholanthrene를 처리하였다. 그 결과 UDP-glucuronosyl-transferase의 mRNA가 3-methylcholanthrene양의 증가에 따라, 처치시간이 길어짐에 따라 증가되어지며 그 mRNA위 크기는 약 2.2Kb 정도임을 알았다. 이로부터 UDP-ghucuronosyltransferase 또한 cytochrome P45O와 함께 다환상 방향족 탄화수소 화합물 조절인자를 통한 조절을 받을 것이며 phase I phase II 약물 대사 효소가 조절상 밀접한 관련을 가짐을 예측할 수 있었다.

• Korean Journal Plant Pathology
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• v.12 no.2
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• pp.182-186
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• 1996

국내에서 재배되고 있는 고추(Capsicum annuum L.)로부터 분리된 TMV pepper 계통을 density gradient centrifugation을 이용하여 순화하였다. 이로부터 바이러스의 total RNA를 분리하였고 RT-PCR에 의하여 TMV pepper 계통의 외피단백질 cDNA를 합성, 증폭하였으며 이를 pBluescript II SK- 벡터에 재조합하였다. 본 실험에서 바이러스 외피단백질과 3` non-coding region을 포함하는 재조합 클론 p1561과 p1562로부터 염기서열을 분석하였고 그 결과로 477 염기의 외피단백질 유전자를 포함하는 691 염기가 합성되었음을 확인하였으며 이것과 TMV common 계통으로부터 합성된 외피단백질 cDNA와의 최대 유사도는 69%였다. 또한 유추된 아미노산 서열에서 이들 두 계통간의 최대 유사도는 81%였다.

• Journal of Life Science
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• v.13 no.5
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• pp.712-717
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• 2003

In order to establish yeast cells that produce leucocin A, a bacteriocin, the 117 bp leucocin A gene with start and stop codons was synthesized and cloned in pAUR123, a yeast vector. Transformed yeast cells showed antibacterial activity against Bacillus subtilis. The leucocin A gene was confirmed by means of PCR methods with plasmid prepared from transformed yeast cells as template and two leucocin A-specific primers. In this results, yeast cells that produce mass amounts of bacteriocin to use as food preservative or antibiotics were established.

• Journal of Industrial Technology
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• v.30 no.B
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• pp.61-68
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• 2010

Three chitinase genes (chiA, chiB, and chiC) were cloned into E. coli by PCR amplification from Serratia marcescens KFRI314. The sizes of cloned chitinase genes were 1692 bp, 1500 bp, and 1443 bp which correspond to 563, 499, and 480 amino acids, respectively. Recombinant chitinases were overexpressed using pHCEIA expression vector and purified to homogenity. The molecular weights of chitinases were about 60kDa, 50 kDa, 52 kDa, respectively. Optimum pHs were around pH 5~6 and optimum temperatures were $45{\sim}50^{\circ}C$ while 90% of enzyme activities were stable up to $50^{\circ}C$. The specific activities of ChiA, ChiB, and ChiC were 233.1, 278.8, $111.3{\mu}mol\;(min)^{-1}\;mg^{-1}$ against colloidal chitin. From experiments using TLC and fluorescent substrate analogues, it was demonstrated that ChiA was endo-chitinase while ChiB and ChiC were chitobiosidase.

• Journal of Sericultural and Entomological Science
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• v.32 no.1
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• pp.44-48
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• 1990

Monoclonal antibodies were prepared against Bombyx densonucleosis virus type-II(Yamanashi isolate). Four hybridoma clones, named C4, Fl, H2, M9 were only reacted with the DNV-II, but those were not reacted with Bombyx densonucleosis virus type-I(Ina isolate) and infectious flacherie virus(IFV) by double diffusion test in 0.8% agarose gel. C4, Fl and M9 of them were reacted with 53KDa polypeptide of DNV-II, and H2 was reacted with 46. 5KDa polypeptide of the virus.

• Microbiology and Biotechnology Letters
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• v.17 no.3
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• pp.178-182
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• 1989

A gene coding for a xylanase of thermophilic alkalophilic Bacillus sp. K-17 was cloned in Escherichia coli C600 with pBR322. Plasmid pAXl13 was isolated from a transformant producing xylanase, and the xylanase gene was located in a 4.3 Kb HindIII fragment. Biotinylated pAXl13 hybridized to a 4.3 Kb HindIII fragment from chromosomal DNA of thermophilic alkalophilic Bacillus sp. K-17. The xylanase activity was observed in the extracellular curture fluid of E. coli carrying pAXl13. The pAXl13-encoded xylanase had the same enzymatic properties as those of xylanase I produced by thermophilic alkalophilic Bacillus sp. K-17.

• Korean Journal of Microbiology
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• v.35 no.1
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• pp.28-34
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• 1999

Fragmentation vectors are used to analyze function and genomic structure of a gene of interest by creating deletion derivatives of large fragments of genomic DNA cloned as yeast artificial chromosomes (YACs). Herein, we developed a new hicistronic fragmentation vector that contains internal ribosomal entry sile (IRES) of encephalomyocarditis vin~s (EMCV) and $\beta$-galactosidase as a reporter gene. This vector system provides a novcl loo1 to analyze expression patterns of a gene of interest due to simultaneous expression of a target gene as well as $\beta$-galactosidase driven from a single message. In addition, the bicistronic fragmentation vector contains four rare-cutting restriction enzyme sites in the polycloning sites which can be used to conveniently insert any kinds of genes and therefore facilitates targeting DNA scgments into YAC by means of homologous recombination. This approach establishes a paradigm for manipulation of mammalian DNA segments and characterization of expression and regulatory regions of mammalian gene cloned as YAC.