• Journal of Microbiology and Biotechnology
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• v.13 no.1
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• pp.110-118
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• 2003

Pseudomonas syringae pv. actinidiae strains, which cause canker disease in kiwifruit, were collected from kiwifruit orchards in Korea and identified using biochemical and physiological tests. The nucleotide sequences of the 16s rDNA and 16s-23s internally transcribed spacer of the isolates were found to be Identical to those of' the pathotype strain, Kwl 1, of P syringae pv. actinidiae. Remarkably, no coding sequence for phaseolotoxin biosynthesis or phaseolotoxin- resistant ornithine carbamoyltransferase was found by PCR amplification in any of the new Korean isolates of pseudomonas syringae pv. actinidiae, although this was clearly identified in the control pathotype Kwl 1 reference strain. In contrast, three primer sets derived from the coronatine biosynthetic gene cluster and DNA from the Korean strains yielded amplified DNA fragments of the expected size. A sequence analysis of the PCR products revealed that P. syringae pv. actinidiae and the Korean strains of pv. actinidiae contain coronafncate ligase genes (cfl)with identical sequences, whereas their. corR genes exhibited 91% sequence similarity. The production of coronatine, instead of phaseolotoxin, by the Korean strains of P. syringae pv. actinidiae was confirmed by a bioassay using reference pathovars known to produce coronatine and phaseolotoxin. The genes for coronatine biosynthesis in the Korean strains of P. syringae pv. actinidiae were found to be present on plasmids.

• Journal of Microbiology
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• v.43 no.2
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• pp.209-212
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• 2005

The purpose of this study was to develop species-specific PCR primers for use in the identification and detection of Actinobacillus actinomycetemcomitans. These primers target variable regions of the 168 ribosomal RNA coding gene (rDNA). We assessed the specificity of the primers against 9 A. actinomycetemcomitans strains and 11 strains (3 species) of the Haemophilus genus. Primer sensitivity was determined by testing serial dilutions of the purified genomic DNAs of A. actinomycetemcomitans ATCC$ 33384^$T Our obtained data revealed that we had obtained species-specific amplicons for all of the tested A. actinomycetemcomitans strains, and that none of these amplicons occurred in any of the other species. Our PCR protocol proved able to detect as little as 4 fg of A. actinomycetemcomitans chromosomal DNA. Our findings suggest that these PCR primers are incredibly sensitive, and should prove suitable for application in epidemiological studies, as well as the diagnosis and monitoring of periodontal pathogens after treatment for periodontitis.

• Fisheries and Aquatic Sciences
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• v.6 no.3
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• pp.116-124
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• 2003

Isolation and cloning of seven-band grouper (Epinephelus septemfasciatus) growth hormone cDNA from pituitary gland revealed an open reading frame of 612 bp coding for a pre-growth hormone of 204 amino acids with a 17 amino acid putative signal peptide. Deduced amino acid sequence showed that there was one possible N-glycosylation site at $Asn^{l84}$ and four cysteine residues $(Cys^{52},\;Cys^{160},\;Cys^{177},\;Cys^{185})$ on t e same positions as in some other species where they were involved in the stabilization of the tertiary structure. The seven-band grouper growth hormone (sbgGH) presented a $99.5\%$ amino acid sequence identity with the growth hormone of Epinephelus coioides and contained the conserved hormone domain region. Comparison of growth hormone sequences from evolutionarily diverse species revealed 25 amino acid residues conserved in jawless fishes to modern mammals. It also revealed an evolutionary trend to retain the same polypeptide sequence even in the distantly related animals while allowing alterations to occur in polypeptides of the closely related species. In order to create a recombinant system to produce high levels of the growth hormone, it was expressed in Escherichia coli (BL21) cells. The gel analysis revealed theoretically expected molecular weights for both mature and pre-sbgGHs.

• Molecules and Cells
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• v.28 no.4
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• pp.341-345
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• 2009

MiRNAs are non-coding RNAs that play a role in the regulation of major processes. The inhibition of miRNAs using antisense oligonucleotides (ASOs) is a unique and effective technique for the characterization and subsequent therapeutic targeting of miRNA function. Recent advances in ASO chemistry have been used to increase both the resistance to nucleases and the target affinity and specificity of these ASOs. Peptide nucleic acids (PNAs) are artificial oligonucleotides constructed on a peptide-like backbone. PNAs have a stronger affinity and greater specificity to DNA or RNA than natural nucleic acids and are resistant to nucleases, which is an essential characteristic for a miRNA inhibitor that will be exposed to serum and cellular nucleases. For increasing cell penetration, PNAs were conjugated with cell penetrating peptides (CPPs) at N-terminal. Among the tested CPPs, Tat-modified peptide-conjugated PNAs have most effective function for miRNA inhibition. PNA-based ASO was more effective miRNA inhibitor than other DNA-based ASOs and did not show cytotoxicity at concentration up to 1,000 nM. The effects of PNA-based ASOs were shown to persist for 9 days. Also, PNA-based ASOs showed considerable stability at storage temperature. These results suggest that PNA-based ASOs are more effective ASOs of miRNA than DNA-based ASOs and PNA-based ASO technology, compared with other technologies used to inhibit miRNA activity can be an effective tool for investigating miRNA functions.

• Journal of Microbiology and Biotechnology
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• v.22 no.2
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• pp.159-165
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• 2012

The rhizobacterial composition varies according to the soil properties. To test if the effect of herbicides on the rhizobacterial communities of genetically modified NK603 glyphosate-tolerant maize varies according to different soil locations, a comparison was made between the effects of glyphosate (Roundup Plus), a post-emergence applied herbicide, and a pre-emergence applied herbicide (GTZ) versus untreated soil. The potential effect was monitored by direct amplification, cloning, and sequencing of the soil DNA encoding 16S rRNA, and high-throughput DNA pyrosequencing of the bacterial DNA coding for the 16S rRNA hypervariable V6 region. The results obtained using three different methods to analyze the herbicide effect on the rhizobacterial communities of genetically modified NK603 maize were comparable to those previously obtained when glyphosate-tolerant maize was grown in soil with different characteristics. Both herbicides decreased the bacterial diversity in the rhizosphere, with Actinobacteria being the taxonomic group most affected. The results suggest that both herbicides affected the structure of the maize rhizobacterial community, but glyphosate was environmentally less aggressive.

• Korean journal of applied entomology
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• v.36 no.1
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• pp.30-36
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• 1997

Except for a cosmopolitan and major pest of apples, Tetranychus urticae Koch, Tetranychus mites in Korea such as T. viennensis Zaher, T. kanzawai Kishida, and T. truncams Ehara have been considered as quarantine pests by Canada and United States. Even though these mites are not feeders on apples, they are suspected to attach accidentally on apple h i t s in autumn as females enter the diapause. The characters used to identify Tetranychus mites have been confined to the shape of aedeagus in adult male. To develope a fast and accurate alternative identification protocol applied to hibernating female mites on apples, their mitochondrial DNA (mtDNA) were examined to find out any polymorphisms to discriminate each species from the other ones. Three pairs of primers for polymerase chain reaction (PCR) were used to amplify cytochrome oxidase subunit I (CO-I) coding region in mitochondrial DNA5 of four species of Tetranycus mites. The longest amplified product was estimated its size as about 680 bp. Digestion with restriction enzymes, AluI, Ddel, and Sau3A, showed length polymorphisms, which will he useful as diagnostic markers to identify Tetranychus mites. Schematic restriction maps in amplified region were shown for each species.

• Korean Journal of Medicinal Crop Science
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• v.14 no.3
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• pp.178-182
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• 2006

This study was conducted to investigate the variation in sequence, the base composition and the sequence similarity of 18S rDNA (18S ribosomal RNA coding region) in the 10 Polygonatum spp. collections. The entire 18S rDNA region of 10 Polygonatum spp. collections ranged from 913 bp to 914 bp. There were 8 variable sites in the entire 18S region, and they were attributable to nucleotide substitution and deletion. $T{\rightarrow}C$ transition happened in 4 sites, and $A{\rightarrow}G$ transition happened in 1 site. $C{\rightarrow}A$ transversion happened in 1 site, and deletion happened in 2 sites. Transition rates were five times that of transversion. Base compositions of 18S rDNA were $23.09{\sim}23.33%$ in adenine, $23.33{\sim}23.52%$ in guanine, $25.60{\sim}25.85%$ in thymine and $27.38{\sim}27.79%$ in cytosine. The A + T content of 18S rDNA of 10 Polygonatum spp. collections averages 48.99%, ranging from 48.80% to 49.18%, and the G + C content averages 51.01%, ranging from 50.82% to 51.20%. Pairwise sequence comparisons indicated that 18S rDNA sequence similarity ranged from 99.7% to 100%.

• Archives of Pharmacal Research
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• v.20 no.5
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• pp.465-470
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• 1997

The human cDNA clone UDPGTh2, encoding a liver UDP-glucuronosyltransferase (UDPGT), was isolated from a .gamma.gt 11 cDNA library by hybridization to mouse transferase cDNA clone, UDPGTm1. The two clones had 74% nlicleotide sequence identities in the coding region. UDPGTh2 encoded a 529 amino acid protein with an amino terminus membrane-insertion signal peptide and a carboxyl terminus membrane-spanning region. In order to establish substrate specificity, the clone was inserted into the pSVL vector (pUDPGTh2) and expressed in COS 1 cells. Sixty potential substrates were tested using cells transfected with pUDPGTh2. The order of relative substrate activity was as follows: 4-hydroxyestrone > estriol >2-hydroxyestriol > 4-hydroxyestradiol > $6{\alpha}$-hydroxyestradiol >$5{\alpha}$-androstane-$3{\alpha}$, $11{\beta}$, $17{\beta}$-triol=5${\beta}$-androstane-$3{\alpha}$ ${\beta}$, $17{\beta}$-triol. There were only trace amounts of gulcuronidation of 2-hydroxyestradiol and 2-hydroxyestrone, and in contrast to other cloned transferase, no gulcuronidation of either the primary estrogens and androgens (estrone, $17{\beta}$estradiol/testosterone, androsterone) or any of the exogenous substrates tested was detected. A lineweaver-Burk plot of the effect of 4-hydroxystrone concentration on the velocity of glucuronidation showed an apparent Km of $13{\mu}M$. The unique specificity of this transferase might play an important role in regulating the level and activity of these potent and active estrogen metabolites.

• Journal of fish pathology
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• v.14 no.2
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• pp.59-64
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• 2001

A complementary DNA encoding metallothionein(MT), a heavy metal-responsive protein was cloned from a cartilaginous shark species. Scyliorhinus torazame. An expressed sequence tag(EST)from the shark liver, which showed high similarity with a MT gene, was isolated and its full-length sequence(390bp)was determined. The putative shark MT cDNA sequence contained an open reading frame consisting 68 amino acids and 182bp of 3-untranslated region including the poly (A+) signal. The deduced amino acid sequence was 41-54% identical to those of other animals including mammals and fish species. Tiger shark MT cDNA showed high conservation in the Cys regions. however, peculiarly contained not only additional five amino acids just prior to the conserved beta-domain but also a Ser residue at C terminal, which has not been seen in other MT sequences.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.17
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• pp.7935-7941
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• 2015

The aim of this study was to implement massive parallel sequencing (MPS) technology in clinical genetics testing. We developed and tested an amplicon-based method for resequencing the BRCA1 and BRCA2 genes on an Illumina MiSeq to identify disease-causing mutations in patients with hereditary breast or ovarian cancer (HBOC). The coding regions of BRCA1 and BRCA2 were resequenced in 96 HBOC patient DNA samples obtained from different sample types: peripheral blood leukocytes, whole blood drops dried on paper, and buccal wash epithelia. A total of 16 random DNA samples were characterized using standard Sanger sequencing and applied to optimize the variant calling process and evaluate the accuracy of the MPS-method. The best bioinformatics workflow included the filtration of variants using GATK with the following cut-offs: variant frequency >14%, coverage ($>25{\times}$) and presence in both the forward and reverse reads. The MPS method had 100% sensitivity and 94.4% specificity. Similar accuracy levels were achieved for DNA obtained from the different sample types. The workflow presented herein requires low amounts of DNA samples (170 ng) and is cost-effective due to the elimination of DNA and PCR product normalization steps.