• 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.

• Molecular & Cellular Toxicology
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• v.4 no.4
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• pp.285-292
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• 2008

Crocin is one of the major components of gardenia fruit and saffron which are widely used as natural food colorants and as traditional Chinese medicines. However, the genotoxicity data on crocin are not sufficient for safety evaluation. The purpose of this study was the examination of the genotoxicity on crocin from gardenia yellow in bacterial and mammalian cells, using various genotoxic battery testing assays and the influence of crocin on methyl methanesulfonate (MMS) and ${H_2}{O_2}$-induced DNA damage in vitro, using single cell gel electrophoresis (comet) assay. From results, no considerable mutagenicity and clastogenicity were seen in bacteria and mammalian cells treated with crocin, by Ames test, chromosomal aberration assay, ${tk}^{+/-}$ gene forward mutation assay and comet assay. And, post-treatment with crocin significantly suppressed ${H_2}{O_2}$-induced DNA damage in a dose-dependent manner. In conclusion, the findings of the present study and other previous observations indicate that crocin has no genotoxic potential. And it showed that crocin clearly repressed the genotoxic potency of ${H_2}{O_2}$. These results suggest that anti-oxidative effects of crocin may be involved in the protective effects of DNA damage.

• Korean Journal of Microbiology
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• v.22 no.3
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• pp.167-173
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• 1984

The cellabiase (${\beta}$-glucosidase) gene in a Cellulomonas sp. CS1-1 was cloned into E. coli HB101 using the vector plasmid pBR322, and the expression of the gene in E. coli studied. The chromosomal DNA of the cellulomonas was digested by seveal restriction enzymes, each of which has only one cleaving site in plasmid pBR322. The recombinant plasmid, pSB2, created with Sal I frament, was expressed for the cellobiase gene in E. coli. The recombiant plasmid was estimated to contain 6.4 Kb foreign DNA at the Sal I site of plasmid pBR322 and the inserted DNA was mapped by single and double digestion with several enzymes. E. coli HB101(pSB2) has slowly grown in a mineral liquid medium containing cellobiose as a sole carbon source. The cellobiase activity in the transformed E. coli was 132 units per liter, which is equivalent to one twenty fifth of that in doner strain Cellulomonas sp. CS1-1. The transforned cell with plasmid containing cellulase gene grow well in the LB mediuns. The synthesis of cellobiase in the strain, E. coli HB101 (pSB2), was inhibited by glucose and at high concentration of cellobiose, and induced by cellobiose at low concentration.

• Journal of Microbiology and Biotechnology
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• v.13 no.2
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• pp.243-250
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• 2003

Eight numerically dominant 4-(2,4-dichlorophenoxy) butyric acid (2,4-DB)-degrading bacteria and three pairs of bacteria showing syntrophic metabolism of 2,4-DB were isolated from soils, and their phylogenetic and phenotypic characteristics were investigated. The isolates were able to utilize 2,4-DB as a sole source of carbon and energy, and their 2.4-DB degradative enzymes were induced by the presence of 2.4-DB. Analysis of 16S rDNA sequences indicated that the isolates were related to members of the genera, Variovorax, Sphingomonas, Bradyrhizobium, and Pseudomonas. The chromosomal DNA patterns of the isolates obtained by polymerase-chain-reaction (PCR) amplification of repetitive extragenic palindromic (REP) sequences were distinct from each other. Four of the isolates had plasmids, but only one strain, DB 1, rad a transmissible 2,4-D degradative plasmid. When analyzed with PCR using primers targeted to the tfdA, B, and C genes, only strains DB2 and DB9a produced DNA bands of the expected sizes with the tfdA and C primers, respectively. All of the isolates were able to degrade 2,4-D as well as 2,4-DB, suggesting that the degradation pathways of these compounds were closely related to each other, but respiratory activities of many isolates adapted to 2,4-DB metabolism were quite low with 2,4-D.

• Journal of Life Science
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• v.12 no.2
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• pp.188-199
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• 2002

A gene coding for xylanase from thermo-tolerant Bacillus pumilus TX703 was cloned into Escherichia coli DH5 $\alpha$ using pUC19. Among 7,400 transformants, four transformants showed clear zones on the detection agar plates containing oat-spells xylan. One of them which showed highest xylanase activity was selected and its recombinant plasmid, named pXES106, was found to carry 2.24 kb insert DNA fragment. When the nucleotide sequence of the cloned xylanase gene (xynK) was determined, xynK gene was found to consist of 1,227 base-pair open reading frame coding for a polypeptide of 409 amino acids with a deduced molecular weight of 48 kDa. The coding sequence was preceded by a putative ribosome binding site, the transcription initiation signals, and cia-acting catabolite responsive element. The deduced amino acids sequence of xylanase is similar to those of the xylanases from Hordeum vulgare (barley) and Clostridium thermocellum, with 39 and 31% identical residues, respectively. The amino acids sequence of this xylanase was quite different from those of the xylanases from other Bacillus species.

• Journal of Ginseng Research
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• v.41 no.4
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• pp.469-476
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• 2017

Background: Panax ginseng Meyer (Asian ginseng) has a large nuclear genome size of > 3.5 Gbp in haploid genome equivalent of 24 chromosomes. Tandem repeats (TRs) occupy significant portions of the genome in many plants and are often found in specific genomic loci, making them a valuable molecular cytogenetic tool in discriminating chromosomes. In an effort to understand the P. ginseng genome structure, we characterized an ultrahigh copy 167-bp TR (Pg167TR) and explored its chromosomal distribution as well as its utility for chromosome identification. Methods: Polymerase chain reaction amplicons of Pg167TR were labeled, along with 5S and 45S rDNA amplicons, using a direct nick-translation method. Direct fluorescence in situ hybridization (FISH) was used to analyze the chromosomal distribution of Pg167TR. Results: Recently, we reported a method of karyotyping the 24 chromosome pairs of P. ginseng using rDNA and DAPI (4',6-diamidino-2-phenylindole) bands. Here, a unique distribution of Pg167TR in all 24 P. ginseng chromosomes was observed, allowing easy identification of individual homologous chromosomes. Additionally, direct labeling of 5S and 45S rDNA probes allowed the identification of two additional 5S rDNA loci not previously reported, enabling the refinement of the P. ginseng karyotype. Conclusion: Identification of individual P. ginseng chromosomes was achieved using Pg167TR-FISH. Chromosome identification is important in understanding the P. ginseng genome structure, and our method will be useful for future integration of genetic linkage maps and genome scaffold anchoring. Additionally, it is a good tool for comparative studies with related species in efforts to understand the evolution of P. ginseng.

• Korean Journal of Medicinal Crop Science
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• v.26 no.2
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• pp.113-118
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• 2018

Background: Gastrodia elata Blume is a saprophytic perennial plant in the Orchidaceae family, because of its agricultural and medicinal effectiveness, researchers focus on its genome and chemical components. However, cytogenetic information based on the chromosome structure and composition to construct chromosomal backbone for genome sequencing research and for the development and breeding of plants is very limited. Methods and Results: We determined the metaphase chromosome composition of the G. elata genome by fluorescence in situ hybridization (FISH) using 5S and 45S rDNAs and telomeric repeat probes. The nuclear genome of G. elata was organized into 2 n = 36, with relatively small ($2.71-5.50{\mu}m$)chromosomes that showed gradual decrease in size. Conglutination phenomenon was observed among the metaphase chromosomes, and it was distinguished from that in other plant metaphase chromosome spreads. One pair of signal was detected for each 5S and 45S rDNA in the pericentromeric region and interstitial region on the short arm of chromosomes 10 and 4, respectively, and telomeric DNA signals were detected in the terminal region of most chromosomes. Conclusions: To our knowledge, this is the first FISH chromosome composition result in G. elata and could be useful in more comprehensive molecular cytogenetic and genomic analyses as well as breeding programs of the medicinal plant G. elata.

• Applied Biological Chemistry
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• v.41 no.3
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• pp.219-223
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• 1998

We have established a transformation system for entomopathogenic fungus, Metarhizium anisopliae, in order to develop mycoinsecticide by recombinant DNA techniques. Protoplasts of M. anisopliae would be transformed to a benomyl-resistant by introducing pBRG-4 plasmid DNA, which contains a ${\beta}-tubulin$ gene of Aspergillus flavus conferring resistance to benomyl and a pyr4 gene of Neurospora crassa, in the presence of 5% polyethylene glycol and 10 mM calcium chloride. Transformants occuring at a frequency of 10 colonies per $50\;{\mu}g$ pBRG-4 DNA grew on the $5\;{\mu}g/ml$ concentrations of benamyl, while the wild type was inhibited by $2.5\;{\mu}g/ml$. From the Southern analysis using genomic DNAs isolated from M. anisopliae transformants, the positive signals suggested that the ${\beta}-tubulin$ gene had integrated in the M. anisopliae genome by homologous recombination.

• Korean Journal of Medicinal Crop Science
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• v.13 no.1
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• pp.48-51
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• 2005

Karyotype analysis and chromosomal locailization of 45S and 5S rDNAs using McFISH (multi-color fluorescence in situ hybridization) were carried out in Jeffersonia dubia Benth., which is one of medicinal plants belonging to Berberidaceae. The somatic metaphase chromosome number was 2n=2x=12 and the size of chromosomes ranged $1.95{\sim}3.50\;{\mu}m$. The chromosome complement consisted of two pairs of metacentrics (chromosomes 1 and 3), two pairs of submetacentrics (chromosomes 2 and 4) and two pairs of subtelocentrics (chromosomes 5 and 6). In McFISH, one pair of 45S rDNA site was detected on the centromeric region of chromosome 2 and three pairs of 5S rDNA sites were detected on the short arm of chromosomes 4, 5 and 6, respectively.

• Journal of Microbiology and Biotechnology
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• v.14 no.2
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• pp.284-289
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• 2004

A PCR assay for the rapid detection of Vibrio vulnificus strains was developed using a virulence gene for elastase found in various Vibrio species. The DNA sequences in the elastase gene facilitated the identification of a species-specific probe for pathogenic V. vulnificus strains from both clinical and environmental sources. Using an elastase gene-based PCR reaction, a species-specific 507-bp PCR product was visualized by agarose gel electrophoresis. Three different DNA extraction methods were then compared to improve the simplicity and rapidity of detection. A PCR assay using the conventional DNA extraction or boiling method was able to detect as few as 25 V. vulnificus cells, making the detection limits at least 1-log-scale lower than that for the EDT A-treated DNA extraction method. In particular, the boiling method, which does not require purification of the chromosomal DNA, was very effective in terms of simple and rapid detection. Meanwhile, the detection limit in a mixed bacterial culture that included other bacteria, such as Escherichia coli or Bacillus subtilis, was two V. vulnificus cells, which was 1-log-scale lower than that for the control. Accordingly, when coupled with a new DNA extraction method, the elastase gene-based PCR can provide a rapid, specific, and sensitive method for identifying V. vulnificus in clinical and environmental samples.