• Title/Summary/Keyword: ITs rDNA

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Distribution Status of Hybrid Types in Large Liver Flukes, Fasciola Species (Digenea: Fasciolidae), from Ruminants and Humans in Vietnam

  • Nguyen, Thi Bich Nga;De, Nguyen Van;Nguyen, Thi Kim Lan;Quang, Huynh Hong;Doan, Huong Thi Thanh;Agatsuma, Takeshi;Le, Thanh Hoa
    • Parasites, Hosts and Diseases
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    • v.56 no.5
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    • pp.453-461
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    • 2018
  • The aim of this study is to delineate 'admixed hybrid' and 'introgressive' Fasciola genotypes present in the Fasciola population in Vietnam. Adult liver flukes collected from ruminants in 18 Provinces were morphologically sorted out by naked eyes for small (S), medium (M) and large (L) body shapes; and human samples (n=14) from patients. Nuclear ribosomal (rDNA) ITS1 and ITS2, and mitochondrial (mtDNA) nad1 markers were used for determination of their genetic status. Total 4,725 worm samples of ruminants were tentatively classified by their size: 6% (n=284) small (S)-, 13% (n=614) medium (M)-, and 81% (n=3,827) large (L)-forms. All the representative (n=120, as 40 each group) and 14 human specimens, possessed maternal mtDNA of only F. gigantica and none of F. hepatica. Paternally, all (100%) of the L-(n=40) and 77.5% (n=31) of the M-flukes had single F. gigantica rDNA indicating 'pure' F. gigantica. A majority (90%, n=36) of the S- and 15% (n=6) of the M-worms had single F. hepatica rDNA, indicating their introgressive; the rest (10%, n=4) of the S- and 7.5% (n=3) of the M-flukes had mixture of both F. gigantica and F. hepatica rDNAs, confirming their admixed hybrid genetic status. Fourteen human samples revealed 9 (64%) of pure F. gigantica, 3 (22%) of introgressive and 2 (14%) of admixed hybrid Fasciola spp. By the present study, it was confirmed that the small worms, which are morphologically identical with F. hepatica, are admixed and/or introgressive hybrids of Fasciola spp., and able to be the pathogens of human fascioliasis.

Effect of Epidermal Growth Factor on Cell Survival of Human Ovarian Teratocarcinoma Cell Line (Epidermal Growth Factor가 난소 기형암종 배아세포주의 생존율에 미치는 효과)

  • Kim, Chung-hui;Kim, Jong-shu;Cuong, Dang Van;Kim, Na-ri;Kim, Eui-yong;Han, Jin
    • Korean Journal of Veterinary Research
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    • v.43 no.2
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    • pp.211-218
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    • 2003
  • Human ovarian cancerous cells survive in a way that they trigger the nucleotide excision repair (NER) or double-strand DNA repair (dsDNA) repair mechanism to show resistance to anticancer drugs and activate many kinds of repair protein, thus removing damaged DNAs. Two experiments on the PA-1 human ovarian teratocareinoma cell line that hardly has any expression of epidermal growth factor receptor (EGFR) were conducted in the study; first, EGF-R was transfected and its receptor was obtained. The receptor was investigated in terms of its mutual relations with many kinds of protein concerning NER or dsDNA repair. Second, it was examined what kind impact cisplatin and adriamycin had on the effects of EGF-R over the PA-1 cell line lacking EGF-R. When being administered with cisplatin and adriamycin, Hey and Hey C2 cell lines showed a high level of resistance while PA-1 cell line a high level of sensitivity. Hey and Hey C2 cell lines that are resistant against anticancer drugs exhibited a high level of EGF-R expression while PA-1 cell line that is sensitive to them did a much lower level of the expression. When PA-1 cell line was transfected for the expression of DNA adduct and EGF-R, it showed a higher level of resistance compared to the control group. There was no difference in the expression of DNA repair proteins (DNA- dependent protein kinase, Ku70, and Ku80) between Hey and the PA-1 cell lines. The results indicate that the Hey cell line that is resistant against cisplatin and adriamycin works along the signaling system responding to the changes of EGF-R while the PA-1 cell line that is sensitive to both of them does to the lack of EGF-R.

Analysis of Cultural Characteristics and Phylogenic Relationships of Collected Strains of Pholiota species

  • Cho, Yong-Hyun;Kong, Won-Sik;Kim, Gyu-Hyun;Jhune, Chang-Sung;You, Chang-Hyun;Yoo, Young-Bok;Kim, Kwang-Ho
    • Mycobiology
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    • v.31 no.4
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    • pp.200-204
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    • 2003
  • Cultural characteristics and phylogenic relationships were investigated and classified among collected strains in Pholiota spp. which contain P. adiposa, P. squarrosa, P. nameko etc. They were tested on the four different media(PDA, MCM, YM, MEA) and sawdust(Alder, Oak, Pine, Popular) substrates. There was a little variation according to the media and sawdust substrates, although PDA and popular sawdust substrate seemed to be better. Most strains showed white colonies, but some strains were brown. Mycelial growth length differed according to the strains. To classify species, the internal transcribed spacer regions(ITS) of the ribosomal DNA(rDNA) repeats from Pholiota spp. were amplified using polymerase chain reaction(PCR) and then sequenced. According to the analysis of ITS sequences, they were classified into five clusters. Their spacer regions were $644{\sim}700$ nucleotides in length. The reciprocal homologies of each ITS region among these strains were ranged from $49.6{\sim}99.9%$. The phylogenic analysis might give a criterion to classify species in the collected strains.

Behavior in Solution and Mixing Ratio-Dependent Binding Modes of Carcinogenic Benzo[a]pyrene-7,8-dione to Calf Thymus DNA

  • Jin, Biao;Han, Sung Wook;Lee, Dong Jin
    • Bulletin of the Korean Chemical Society
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    • v.35 no.10
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    • pp.3015-3020
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    • 2014
  • The behavior of benzo[a]pyrene-7,8-dione (BPQ) in aqueous solution and its interaction with native DNA was investigated using conventional absorption and linear dichroism (LD) spectroscopy. The appearance of a broad absorption maximum at long wavelengths and its proportional relationship to solvent polarizability suggested that BPQ adopts a aggregated state for all solutions examined. Disappearance of this absorption band at higher temperatures in aqueous solution also supported BPQ aggregation. When associated with DNA absorption spectral properties were essentially the same as that in aqueous solution. However, two isosbestic wavelengths were found in the concentration-dependent absorption spectrum of the BPQ-DNA complex, suggesting the presence of at least two or more DNA-bound BPQ species. Both species produced $LD^r$ spectra whose magnitude in BPQ absorption region is larger or comparable to that in the DNA absorption region, suggesting that the molecular BPQ plane is near perpendicular relative to the local DNA helical axis. Therefore, BPQ molecules are aligned along the DNA stem in both DNA-aggregated BPQ species.

Identification and Characterization of an R-M System in Paracoccus denitrifican DYTN-1 to Improve the Plasmid Conjugation Transfer Efficiency

  • Yunpeng Shi;Wenyan Cao;Zhiping Zheng;Sha Xu;Lijuan Chai;Shenghu Zhou;Yu Deng
    • Journal of Microbiology and Biotechnology
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    • v.34 no.9
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    • pp.1826-1835
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    • 2024
  • Paracoccus denitrificans has been identified as a representative strain with heterotrophic nitrificationaerobic denitrification capabilities (HN-AD), and demonstrates strong denitrification proficiency. Previously, we isolated the DYTN-1 strain from activated sludge, and it has showcased remarkable nitrogen removal abilities and genetic editability, which positions P. denitrificans DYTN-1 as a promising chassis cell for synthetic biology engineering, with versatile pollutant degradation capabilities. However, the strain's low stability in plasmid conjugation transfer efficiency (PCTE) hampers gene editing efficacy, and is attributed to its restriction modification system (R-M system). To overcome this limitation, we characterized the R-M system in P. denitrificans DYTN-1 and identified a DNA endonuclease and 13 DNA methylases, with the DNA endonuclease identified as HNH endonuclease. Subsequently, we developed a plasmid artificial modification approach to enhance conjugation transfer efficiency, which resulted in a remarkable 44-fold improvement in single colony production. This was accompanied by an increase in the frequency of positive colonies from 33.3% to 100%. Simultaneously, we cloned, expressed, and characterized the speculative HNH endonuclease capable of degrading unmethylated DNA at 30℃ without specific cutting site preference. Notably, the impact of DNA methylase M9 modification on the plasmid was discovered, significantly impeding the cutting efficiency of the HNH endonuclease. This revelation unveils a novel R-M system in P. denitrificans and sheds light on protective mechanisms employed against exogenous DNA invasion. These findings pave the way for future engineering endeavors aimed at enhancing the DNA editability of P. denitrificans.

PDMS/Glass Based DNA Microbiochip for Restriction Enzyme Reaction and Electrophoresis Detection (DNA의 제한효소 반응 및 전기영동 검출용 PDMS/유리 마이크로바이오칩)

  • Choi Joon-Young;Ahn Yoo Min;Hwang Seung-Yong
    • Transactions of the Korean Society of Mechanical Engineers A
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    • v.30 no.1 s.244
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    • pp.26-31
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    • 2006
  • This paper reports low-cost PDMS/glass based DNA microbiochip for the restriction enzyme reaction and its products detection using the capillary electrophoresis. The microbiochip ($25mm{\times}75mm$) has the heater integrated reactor ($5{\mu}{\ell}$) for DNA restriction enzyme reaction at $37^{\circ}C$ and the microchannel ($80\;{\mu}m{\times}100\;{\mu}m{\times}58mm$) for the capillary electrophoresis detection. It is experimentally confirmed that the digestion of the plasmid ($pGEM^{(R)}-4Z$) by the enzyme (Hind III and Sca I) is performed for less than 10 min and its electrophoresis detection is able to sequentially on the fabricated microbiochip.

Karyotype Analysis and Physical Mapping of rDNAs Using Bicolor-FISH in Tiarella polyphylla D. Don (헐떡이풀의 핵형분석과 Bicolor FISH를 이용한 물리적 지도 작성)

  • Kim, Soo-Young;Lee, Joong-Ku
    • Korean Journal of Plant Resources
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    • v.20 no.5
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    • pp.446-450
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    • 2007
  • Tiarella polyphylla D. Don(Saxifragaceae) is a perennial herb and distributed in China, Japan, Taiwan and Korea. Especially, it only grows in Ulleung island of Korea. It has been using for asthma, bruise and audition troubles with main components of some Triterpenoids and seven oleanolic Saponins. There is only known its chromosomal number rarely and cytogenetic study was not done. From this study, karyotype analysis and chromosomal localization of 5S and 45S rDNAs using bicolor-FISH(fluorescence in situ hybridization) were carried out. The somatic metaphase chromosome number was 2n=2x=14 and the size of chromosomes ranged $1.66{\sim}3.50{\mu}m$. The chromosome complement consisted of four pairs of submetacentrics(chromosomes 1, 2, 3 and 6), two pairs of subtelocentrics(chromosomes 5 and 7) and one pair of telocentrics(chromosome 4). We also observed NOR(nucleolus organizer region) on the chromosome 4. In bicolor-FISH, one pair of 55 and 45S rDNA sites was detected on the centromeric region of chromosome 3 and short arm of chromosome 4, respectively. Bicolor FISH was very useful tool for the localization and identification of rDNAs on the chromosomes in Tiarella polyphylla.

Development of Molecular Biological Methods to Analyze Bacterial Species Diversity in Freshwater and Soil Ecosystems

  • Lee, Dong-Hun;Noh, Sung-Ae;Kim, Chi-Kyung
    • Journal of Microbiology
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    • v.38 no.1
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    • pp.11-17
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    • 2000
  • A new method was developed for the rapid analysis of diverse bacterial species in the natural environment. Our method is based on PCR-single-strands-conformation polymorphism (PCR-SSCP) and selective isolation technique of single-stranded DNA. Variable V3 fragments of 16S rDNA were amplified by PCR with bacterial 16S rDNA primers, where one of the primers was biotinylated at the 5'-end. The biotinylated strands of the PCR products were selectively isolated by using streptavidin paramagnetic particles and a magnetic stand, to prevent SSCP analysis producing heteroduplexes from heterogeneous DNA samples. The selected strands were separated by electrophoresis on a polyacrylamide gel, and detected by silver staining. Analysis of PCR products from 8 bacterial strains demonstrated their characteristic DNA band patterns. In addition, changes in the structure of the bacterial community and species diversity in the microcosm treated with phenol could be monitored. After 3 weeks of incubation, phenol and its intermediate, 2-hydroxy-muconic-semialdehyde, were degraded by indigenous bacteria. These dominating bacterial populations were identified as strong bands on an SSCP gel. Therefore, this study provides useful tools for microbial community analysis of natural habitats.

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Three Intraspecific groups in Korean Isolates of Phytophthora drechsleri Based on PCR-RFLP of Ribosomal DNA (Ribosomal DNA의 PCR-RFLP에 의한 국내산 Phytophthora drechsleri의 3가지 종내그룹)

  • 홍승범;지형진;이승임;고승주;류진창;김인수
    • Korean Journal Plant Pathology
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    • v.14 no.5
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    • pp.519-525
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    • 1998
  • Intraspecific genetic diversity of Korean isolates of Phytophthora drechsleri was investigated based on PCR-RFLP of rDNA along with closely related species in the genus; P. cryptogea, P. melonis, P. erythroseptica, P. cinnamomi, P. cambivora and P. cactorum. Gene regions of nuclear small subunit and internal transcribed spacer (ITS) in rDNA were amplified with polymerase chain reaction and digested with 9 restriction enzymes. Phytophthora species was readily differentiated from each other based on the digestion patterns, however, P. cryptogea was not separable from some isolates of P. drechsleri. Twenty one isolates of P. drechsleri originated from 15 host plants were divided into three distinct groups designated as PdG1, PdG2 and PdG3, respectively. Four isolates in PdG1 were originated from green vegetables and tomato and nine isolates in PdG2 were mainly isolated from medicinal plants. The two groups showed 95.3% homology and four isolates of P. cyptogea came under the groups. However, Eight isolates in PdG3 collected from cucurbits were clearly differentiated from those of PdG1 and PdG2 by 66.5% homology, but completely matched with a Taiwan isolate of P. melonis. Results indicated that three distinct groups exist in Korean isolates of P. drechleri and each group has host preference. In addition, reclassification of the cucurbits isolates are reserved because of their distinct genetic characters from other intraspecific groups in P. drechsleri.

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