• Journal of Microbiology and Biotechnology
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• v.2 no.2
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• pp.115-121
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• 1992

In order to broaden the spectrum of substrate utilization of a Gram negative bacterium Zymomonas mobilis which has a great potential as an industrial ethanol producing microorganism, cloning of $\alpha$-amylase gene into Z. mobilis ZM4 was tried. The $\alpha$-amylase gene was isolated from Bacillus stearothermophilus. By Southern blot analysis, it was proven that the $\alpha$-amylase gene fragment was originated from a naturally occuring plasmid of B. stearothermophilus ATCC 31195. To place $\alpha$-amylase gene under the control of Z. mobilis promoter, two different Z. mobilis expression vectors, pZA26 and pLOI204, were used. The truncated $\alpha$-amylase gene was then introduced into these vectors. Both qualitative and quantitative activities of $\alpha$-amylase were observed in Z. mobilis cells harboring these plasmids with the $\alpha$-amylase gene inserted. Gas chromatographic analysis of ethanol showed that one of the Z. mobilis transconjugants was capable of producing 67 mM ethanol from rich medium(RM) containing 5% soluble starch as a sole carbon source.

• Journal of Ginseng Research
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• v.19 no.1
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• pp.51-55
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• 1995

The DNA fragment containing ginseng ribulose-1,5-bisphosphate carboxytase/oxygenase large subunit(rbcL) gene was cloned from the ginseng chloroplast EcoRl library by colony lift hybridization with tobacco rbcL gene probe. From the screened clone, the DNA fragment containing ginseng rbcL gene was digested with several restriction enzyme and analyzed by Southern blot hybridization for the construction of restriction map. The ginseng rbcL gene fragment was subcloned in pBluescript II SK + vector and sequence analysis was performed. The nucleotide sequence of ginseng rbcL gene was compared with those of petunia, tobacco, alfalfa, rice and barley, which showed a homology of 93.1%, 95.2%, 90.5%, 85.5% and 84.3%, respectively.

• Microbiology and Biotechnology Letters
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• v.14 no.3
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• pp.213-218
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• 1986

Hybrid plasmid pEA24, shuttle vector YRp7 carrying amylase gene of Bacillus amyloliquefaciens, was transformed to yeast Saccharomyces cerevisiae, and the expression of B. amyloliquefaciens amylase gene in yeast was investigated. The frequency of transformation to S. cerevisiae DBY747 with YRp7 was increased by treatment of 40% polyethylene glycol (MW 4, 000), PH 7.0, at 3$0^{\circ}C$, and by regeneration used 2% top agar. The amount of cellular amylase activity produced by S. cerevisiae containing pEA24 was 2% of that secreted from B. amyloliquefaciens, but in case of S. cerevisiae transformant, the amylase secreted was not detected. A comparison of genetic stability of pEA24 and YRp7 plasmids in yeast was carried out by cultivation of transformants in tryptophan-supplement-medium. The pEA24 plasmid was more unstable than YRp7 in S. cerevisiae. The size of pEA24 extracted from S. cerevisiae transformants was found to be identical with that from E. coli transformants by agarose gel electrophoresis.

• 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.30 no.3
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• pp.149-153
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• 1992

Cloning of the gene encoding extracellular .betha.-lactamase from Streptomyces sp. SMF13 in a plasmid pIJ702 and expression of the gene in Streptomyces invidans were carried out. Optimal conditions for the formation of protoplasts of S.lividans and the regeneration of the protoplasts were evaluated. Streptomyces sp. SMF-13 was selected as a donor strain of .betha.-lactamase gene and totla DNA of the strain was partially digested with Sau3A I. DNA fragments ranged from 4kb to 10 kb were ligated to pIJ702 AT Bgl II site and then the ligated DNAs were transformed to the protoplasts of S, livivans. The transformation efficiency was $2 *10^{3}$ .$\mu$g DNA for the ligated DNA mixture. One colony among a thousand colonies regenerated showed extracellular .betha.-lactamase and the size of the inserted DNA fragment was estimated to be 3.94 kb. The .betha.-lactamase activity in the culture broth of the recombinant strain was maximum at 3 days culture to be 1.0 unit/ml.

• Journal of Microbiology and Biotechnology
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• v.10 no.1
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• pp.86-90
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• 2000

We have earlier reported on the cloning and identification of bft-k from an enterotoxigenic strain of Bacteroides fragilis 419, which was isolated from the blood of a Korean patient who suffered from systemic infections [4,5]. The bft-k gene encodes a 397-amino-acids metalloprotease enterotoxin, and the protein has been identified as a new isoform of B. fragilis enterotoxins (BFTs), which are cytopathic to intestinal epithelial cells to induce fluid secretion and tissue damage in ligated intestinal loops [4, 6, 18, 20]. This report describes the cloning and sequencing of the enterotoxin pahogenicity islet of B. fragilis 419 which contains the bft-k gene. the cloned enterotoxin pathogenicity islet was found to have 6,045 bp in length and to contain 120bp direct repeats near its end. In the pathogenicity islet, in addition to the BFR-K, two putative open reading frames (ORFs) were identified; (1) the t-3 gene encoding a 396-amino-acids protein of a putative metalloprotease; (2) the third gene encoding an ORF of a 59-amino-acids protein, whose function has not yet beenn characterized. The expression of the t-3 gene in B. fragilis 419 was verified by western blot analysis.

• Microbiology and Biotechnology Letters
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• v.17 no.2
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• pp.160-164
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• 1989

A 5.7Kb EcoRI fragment containing alkaline amylase gene of Bacillus sp. AL-8 obtained in the previons experiment (10) was transformed in B. subtilis via plasmid pUB110. The enzymatic proper-ties of the amylase produced by the transformants were Identical to those of the donor strain. Thus, the alkaline amylase activity from the transformant was maximum at pH 10 and 5$0^{\circ}C$. And the enzyme was very stable over the ranges of alkaline pH. In order to determine the location of the alkaline amylase gene within the 5.7Kb DNA fragment, the fragment was subcloned in E. coli. It was found that the alkaline amylase gene was located k EcoRI fragment of 3.7Kb.

• The Korean Journal of Mycology
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• v.17 no.1
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• pp.35-38
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• 1989

Gene libraries of DNA from Flammulina velutipes were constructed using Escherichia coli plasmid pBR 322. Leu 2 gene coding ${\beta}-isopropylmalate$ dehydrogenase from F. velutipes was cloned by complementation of leucine requiring mutant of E. coli. The size of inserted DNA fragment of this clone is about 1 Kbp. The fragment has Bam H1 and Ava 1 restriction sites.

• Reproductive and Developmental Biology
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• v.29 no.1
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• pp.15-18
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• 2005

This study was carried out to analyze the structure of the gene related to porcine stress syndrome (PSS) through cloning from PSS heterozygous pigs and to examine hereditary type associated with PSS. The results obtained from the study were summarized as follows: Amino acid arginine and cysteine exist in dominant gene, N and harmfully recessive gene, n, respectively. It was confirmed that such genes exists as alleles in the sixth chromosome related to PSS. These allelic genes might be inherited according to Mendelian law.

• 한국생물공학회:학술대회논문집
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• 2000.04a
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• pp.153-156
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• 2000

Xylan, the major hemicellulose component of many plants, occurs naturally in a partially acetylated form and lignin, the most resistant component in plant cell wall degradation, is also attached to ${\beta}-1,4-linked-D-xylose$ backbone through the ester linkage. Esterases are required to release the esterified substituent and acetyl esterases are important in the complete degradation of acetylated polysaccharides, like pectins and xylans. The gene(Axe) encoding acetyl xylan estarase(AXE) was isolated from genomic ${\lambda}$ library from Aspergillus ficuum. Nucleotide sequencing of the Axe gene indicated that the gene was separated with two intervening sequences and the amino acid sequence comparison revealed that it was closely related to that from A. awamori with the 92 % indentity. Heterologous expression of AXE was conducted by using YEp352 and Saccharomyces cerevisae 2805 as a vector and host expression system, respectively. The Axe gene was placed between GAL1 promoter and GAL7 terminator and then this recombinant vector was used to transform S. cerevisiae 2805 strain. Culture filtrate of the transformed yeast was assayed for the presence of AXE activity by spectrophotometry and, comparing with the host strain, four to five times of enzyme activity was detected in culture filtrate of transformed yeast.