• Journal of Life Science
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• v.23 no.2
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• pp.290-298
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• 2013

The genus Mycobacterium includes crucial animal and human pathogens such as Mycobacterium tuberculosis, Mycobacterium leprae, and Mycobacterium bovis. Although it is important to understand the genetic basis for their virulence and persistence in host, genetic analysis in mycobacteria was hampered by a lack of sufficient genetic tools. Therefore, many functional vectors as molecular genetic tools have been designed for understanding mycobacterial biology, and the application of these tools to mycobacteria has accelerated the study of mechanisms involved in virulence and gene expression. To overcome the pre-existing problems in genetic manipulation of mycobacteria, this paper reports new vector systems as effective genetic tools in Mycobacterium smegmatis. Three vectors were developed; pKOTs is a suicide vector for mutagenesis containing a temperature-sensitive replication origin (TSRO) and the sacB gene encoding levansucrase as a counterselectable marker. pMV306lacZ is an integrative lacZ transcriptional fusion vector that can be inserted into chromosomal DNA by site-specific recombination. pTnMod-OKmTs is a minitransposon vector harboring the TSRO that can be used in random mutagenesis. It was demonstrated in this study that these vectors effectively worked in M. smegmatis. The vector systems reported here are expected to successfully applicable to future research of mycobacterial molecular genetics.

• Asian-Australasian Journal of Animal Sciences
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• v.21 no.1
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• pp.11-18
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• 2008

We created a cDNA microarray representing approximately 3,500 pig genes for functional genomic studies. The array elements were selected from 6,494 cDNA clones identified in a large-scale expressed sequence tag (EST) project. These cDNA clones came from normalized and subtracted porcine adipose tissue cDNA libraries. Sequence similarity searches of the 3,426 ESTs represented on the array using BLASTN identified 2,790 (81.4%) as putative human orthologs, with the remainder consisting of "novel" genes or highly divergent orthologs. We used the gene microarray to profile transcripts expressed by adipose tissue of fatty Chinese Xiang pig (XP) and muscley Large White (LW). Microarray analysis of RNA extracted from adipose tissue of fatty XP and muscley LW identified 81 genes that were differently expressed two fold or more. Transcriptional differences of four of these genes, adipocyte fatty acid binding protein (aP2), stearyl-CoA desaturase (SCD), sterol regulatory element binding transcription factor 1 (SREBF1) and lipoprotein lipase (LPL) were confirmed using SYBR Green quantitative RT-PCR technology. Our results showed that high expression of SCD and SREBF1 may be one of the reasons that larger fat deposits are observed in the XP. In addition, our findings also illustrate the potential power of microarrays for understanding the molecular mechanisms of porcine development, disease resistance, nutrition, fertility and production traits.

• Fisheries and Aquatic Sciences
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• v.20 no.8
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• pp.16.1-16.8
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• 2017

The exportation of cultured olive flounder (Paralichthys olivaceus) in Korea has been recently decreasing due to the infections with a myxozoan parasite Kudoa septempunctata, and there is a strong demand for strict food safety management because the food poisoning associated with consumption of raw olive flounder harbouring K. septempunctata has been frequently reported in Japan. The life cycle and infection dynamics of K. septempunctata in aquatic environment are currently unknown, which hamper establishment of effective control methods. We investigated sea water and marine invertebrates collected from olive flounder farms for detecting K. septempunctata by DNA-based analysis, to elucidate infection dynamics of K. septempunctata in aquaculture farms. In addition, live marine polychaetes were collected and maintained in well plates to find any possible actinosporean state of K. septempunctata. The level of K. septempunctata DNA in rearing water fluctuated during the sampling period but the DNA was not detected in summer (June-July in farm A and August in farm B). K. septempunctata DNA was also detected in the polychaetes Naineris laevigata intestinal samples, showing decreased pattern of 40 to 0%. No actinosporean stage of K. septempunctata was observed in the polychaetes by microscopy. The absence of K. septempunctata DNA in rearing water of fish farm and the polychaetes N. laevigata intestinal samples during late spring and early summer indicate that the infection may not occur during this period. N. laevigata was suspected as the possible alternate invertebrate host of K. septempunctata, but the actinosporean stage was not found by well plate method and further studies will be necessary. This research provides important baseline information for understanding the infection dynamics of K. septempunctata in olive flounder farms and further establishment of control strategies.

• Korean Journal of Veterinary Service
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• v.41 no.4
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• pp.257-262
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• 2018

Avian pathogenic E. coli (APEC) causes severe economic losses in the poultry farms, due to systemic infections leading to lethal colisepticemia. It causes a variety of diseases from air sac infection to systemic spread leading to septicemia. Secondary infection contains opportunistic infections due to immunosuppression disease. Collibacillosis causes the great problems in the poultry industry in Korea. Thus, it is necessary to identify and classify the characteristics of E. coli isolate of chicken origin to confirm the diversity of symptoms and whether they are transmitted among the farms. Fragment analysis is identify the difference in the number of Variable-Number Tandem-Repeats (VNTRs) for genotyping. VNTRs have repeating structure (Microsatellite, Short tandem repeats; STR, Simple sequence repeats; SSR) in the chromosome. This region can be used as a genetic marker because of its high mutation rate. And various lengths of the amplified DNA fragment cause the difference in the number of repetition of the DNA specific site. The number of repetition sequences indicates the separated size of fragments, so the each fragments can be distinguished by specific samples. The results of the sample show that there is no difference in six microsatellite loci (yjiD, aidB, molR_1, ftsZ, b1668, yibA). There are differences among the farms in relation of the number of repetitions of other six microsatellite loci (ycgW, yaiN, yiaB, mhpR, b0829, caiF). Four (ycgW, yiaB, b0829, caiF) of these six microsatellite loci show statistically significant differences (P<0.05). It means that the analysis using four microsatellite loci including ycgW, yiaB, b0829, and caiF can confirm among the farms. Five E. coli samples in one farm have same SSR repetition at all markers. But, there are significant differences from other farms at Four (ycgW, yiaB, b0829, caiF) microsatellite loci. These results emphasize again that the four microsatellite loci makes a difference in the amplified DNA fragments, enabling it to be used for E. coli genotyping.

• Journal of Life Science
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• v.16 no.1
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• pp.30-36
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• 2006

Neutrophils are short-lived leukocytes that play a vital role in immune responses to bacteria, yeast, and fungi. This study was performed to investigate the effect of 4-O-methyl-ascochlorin (MAC), an anti-tumor, antibiotic, and anti-fungal prenyl-phenol compound on the spontaneous apoptosis of human neutrophils. MAC time- and dose-dependently accelerated the spontaneous apoptosis of human neutrophils. The effect of MAC on neutrophil apoptosis was blocked by pre-treatment of the neutrophils with specific inhibitors of pancaspase (zVAD-fmk), caspase-8 (zIETD-fmk), or caspase-3 (zDEVD-fmk). The cleavage of procaspase-8 and procaspase-3 was increased by MAC. Mitochondrial permeability, which was measured by the retention of $DiOC_6(3)$, was dose-de-pendently increased by MAC but the change of mitochondrial permeability was not blocked by pretreatment of neutrophils with zIETD-fmk. These results suggest that MAC induces neutrophil apoptosis by caspase-8-dependent but mitochondria-independent manner.

• Clinical and Experimental Pediatrics
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• v.45 no.10
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• pp.1263-1272
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• 2002

Purpose : Rett syndrome(RTT) is an X-linked dominant neurodevelopmental disorder affecting 1 per 10,000-15,000 female births worldwide. It was initially described by Andreas Rett in 1966. RTT involves developmental regression characterized stereotypic hand movements, tremors, gait apraxia, seizures, deceleration of head growth after the age of 6-18 months. The disease-causing gene was identified as MECP2 on chromosome Xq28. We carried out mutational analysis of MECP2 genes in RTT patients. Methods : Whole blood(5 cc) of 34 sporadic RTT patients was collected in EDTA-anticoagulated tubes. Genomic DNA was extracted from peripheral blood using the E.Z.N.A. blood DNA kit. Four exons of the MECP2 gene were amplified by PCR in 34 Korean with RTT. We carried out PCR divided the exon three into two parts and the exon four into five parts. Primer sequences designed by Amir et al. in 1999 were almost used(AF030876). Sequencing primers used were the same as PCR. DNA sequencing reactions were performed using an ABI 377 DNA sequencer and ABI PRISM dye terminator cycle sequencing reaction kit(Perkin-elmer). The results were compared with the normal DNA sequence(X99686). To confirm the change of sequence on novel mutations, RFLP analysis was performed. Results : The MECP2 mutations were detected in 23(67.6%) of the 34 patients. The mutations consisted of 12 different types including nine missense and three nonsense mutations. Of these, three (L100V, G161E and T311M) mutations were newly identified. Most of the mutations discovered are located within MBD(39.1%) and TRD(39.1%). In this study, three(T158M, R270X, R306C) mutations were identified high frequency. Conclusion : MECP2 gene was also an important cause of Korean RTT patients. MECP2 gene study is an important tool for diagnosis of Korean RTT patients.

• Korean Journal of Microbiology
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• v.33 no.2
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• pp.149-156
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• 1997

We have employed comparative RFL,P(Restriction Fragment Ixngth Pol~iniorphism) analysis and molecular phylogenetic techniques to investigate the diversity of uncultured microorganisms associated with the anaerobic PCP degradation in PCP-adapted enrichment cultures inoculated by samples from anaerobic cewage sludgc(Jangrim, Pusan) and leachate of landfill site(Kimhae). 16s rDNA cloncs were obtairted by PCR amplification of mixed population DNAs extracted directly from the nonactive and active stage ol each PCP-adapted culture. After three rounds of comparative RFLP analyses. two RFLP types. designated as Ala and Hld, were found prevalent and common in both active stage samples. Thc analysis of phylogenctic diversity bawd on the 5'-terminal 180 nt of sequences from whole clones of the Ala and Bld RFLP types showed close similarity among themselves. In case of Bld clones, 7XQ of them shared identical sequences. Thcse resuliq suggest that the clones of both RFLP types wcre originated from highly affiliated microorganisms which are e~iriched as a result of metabolic activity to PCP. The full-length 16s rRNA sequence of each representative clone from both RFLP types was determined. and an Ala clone w i n found to he related to Clo.strrdiurn ulfutzac~(Genk~ank No. Z69203) and a Bld clone to Thermobacteroides proteolyticus(Genbank No. X09335), with sequence similarities of 89%' and 97%. respectively.

• Preventive Nutrition and Food Science
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• v.5 no.3
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• pp.153-159
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• 2000

A 4.4 kb DNA fragment encompassing lacA (galactoside acetyltransferase) and lacZ($\beta$-galactosidase) genes from Lactococus lactis ssp. lactis ATCC 7962 (L. lactis 7962) was introduced ito a Lac strain, Lactococcus lactis ssp. lactis MG1363 (L. lactis MG1363) by using a lactococcal expression vector, pMG36e and expression level of lacZ was examined. Growth rates and $\beta$-galactosidase ($\beta$-gal) activities of MG1363 cells carrying recombinant plasmid, pMLZ3, on M17 broth containing different carbon sources (1%, w/v) were examined. Contrary to the expectations, MG1363 [pMLZ3] grown on lactose showed the lowest enzyme activity (17 units) and cells grown on galactose had the highest $\beta$-gal activity (41 units). Cells grown on glucose had intermediate activity (33 units). These activities are about one tenth of the values observed in L. lactis 7962 where lacZ is present as a single-copy gene in the chromosome. When the cellular concentrations of lacZ transcript were examined using slot blot hybridization, it was found that MG1363[pMLZ3] produced sufficient amounts of transcript. These results indicate that either proteolytic degradation of $\beta$-gal or other regulatory mechanism prevent the translation or accumulation of $\beta$-gal in L. lactis MG1363 cells. In regard to regulation, the presence of the ccpA gene in L. lactis MG1363 was confirmed by Southern blot.

• Korean Journal of Environmental Biology
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• v.17 no.4
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• pp.503-512
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• 1999

The morphological variations and habitat environment of the Zostera asiatica Miki (Zosteraceae) were examined in the waters off the eastern coast of Korea and Akkeshi Bay of Japan. In this study, we report new habitat of Z. asiatica along the eastern coast of south Korea； Gonghunjin, Donghori, Youngdong and Chilpo. Z. asiatica is found at the depth of 8∼15 m in these areas. However, Z. asiatica grows at the depth of 3 m in Akkeshi Bay of Japan. The sediment of habitats in the eastern coast of Korea were principally composed of sand. The stems were 64.0∼75.7 cm long in mature plants with flowering shoots and the leaves were 11.0∼15.0 mm wide for Korean populations. Plants from Akkeshi Bay of Japan had stems of 187.0∼234.0 cm in length. The difference in size are considered to be due to the depth of habitats. In spite of the significant morphological difference between Korea and Japan population of Z. asiatica, there was no difference in ITS sequences.

• Journal of Aquaculture
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• v.7 no.1
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• pp.41-54
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• 1994

In order to evaluate the availability of lacZ as a reporter gene for producing transgenic mud loach, foreign DNA, bacterial \beta-galactosidase$ gene (lacZ) was microinjected into mud loach eggs and its insertion and expression were examined. X-gal based histochemical assay, fluorimetric analysis of \beta-galactosidase$ with 4-methylumbelliferyl-$\beta$-D-galactoside (MUG) and molecular biological examination using polymerase chain reaction (PCR), dot blot, southern blot and sequence analysis of PCR products were carried out to analyze both microinjected group and non-injected controls. The results are disccussed in this paper.