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Multiplex PCR Using Conserved and Species-Specific 16S rDNA Primers for Simultaneous Detection of Fusobacterium nucleatum and Actinobacillus actinomycetemcomitans

  • Kim, Mi-Kwang;Kim, Hwa-Sook;Kim, Byung-Ock;Yoo, So-Young;Seong, Jin-Hyo;Kim, Dong-Kie;Lee, Shee-Eun;Choe, Son-Jin;Park, Joo-Cheol;Min, Byung-Moo;Jeong, Moon-Jin;Kim, Do-Kyung;Shin, Yong-Kook;Kook, Joong-Ki
    • Journal of Microbiology and Biotechnology
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    • v.14 no.1
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    • pp.110-115
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    • 2004
  • This study was undertaken to develop PCR primers for the simultaneous detection of Fusobacterium nucleatum and Actinobacillus actinomycetemcomitans, using two species-specific reverse primers in combination with a single conserved forward primer. These primers target the variable and conserved regions of the 16S rDNA. The primer specificity was tested against (i) four F. nucleatum and three A. actinomycetemcomitans strains and (ii) seven representatives of the different species of oral bacteria. The primer sensitivity was determined by testing serial dilutions of the purified genomic DNA of F. nucleatum and A. actinomycetemcomitans. The data indicate that species-specific amplicons could be obtained for all the F. nucleatum and A. actinomycetemcomitans strains tested, which were not found in the seven other species. The multiplex PCR could detect as little as 4 fg of chromosomal DNA of F. nucleatum and A. actinomycetemcomitans simultaneously. These findings suggest that these PCR primers are highly sensitive and are suitable for applications in epidemiological studies, diagnosis, and monitoring F. nucleatum and A. actinomycetemcomitans after the treatment of periodontitis.

Enact of Glutathione(GSH/GSSG) Contents of Fermented Ginseng on the ${\gamma}$-irradiated Liver of Mice (감마선을 조사한 생쥐 간에서 발효인삼이 Glutathione 함량에 미치는 영향)

  • Ko, In-Ho
    • The Journal of Korean Society for Radiation Therapy
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    • v.18 no.1
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    • pp.29-34
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    • 2006
  • Purpose: The radioprotective effects of white and fermented ginseng on liver damage induced by $^{60}Co\;{\gamma}$-ray were investigated. Materials and Methods: To one group of ICR male mice were given white(150 mg/kg/day for 7 days, orally) and fermented ginseng(150 mg/kg/day for 7 days, orally) before $^{60}Co\;{\gamma}$-ray irradiation. To another group were irradiated by 5 Gy(1.01 Gy/min) dose of $^{60}Co\;{\gamma}$-ray. Contrast group were given with saline(0.1 mL). The levels of reduced(GSH) and oxidized(GSSG) glutathione in liver tissue were measured. Results: In the fermented(150 mg/kg) and white ginseng(150 mg/kg) groups than irradiation group, the GSH levels were significantly increased, but the GSSG levels were significantly decreased. The ratio of GSSG/total GSH was significantly decreased in the fermented(150 mg/kg) and white ginseng(150 mg/kg) groups than irradiation group. Conclusion: In the fermented(150 mg/kg) groups than white ginseng(150 mg/kg) groups the GSH levels were significantly increased. The radioprotective effects of fermented(150 mg/kg) groups than white ginseng(150 mg/kg) groups were increased.

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DNA Sequence analysis and rfbM gene amplification using PCR for detect salmonella C1 serogroup (살모넬라 C1 serogroup 특이 rfbM 유전자 증폭과 염기서열 분석)

  • Lee, Sung-il;Jung, Suk-chan;Moon, Jin-san;Park, Yong-ho;Lee, John-wha;Kim, Byeong-su;Baek, Byeong-kirl
    • Korean Journal of Veterinary Research
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    • v.36 no.1
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    • pp.109-118
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    • 1996
  • The Salmonella rfb gene encoding for the biosynthesis of the oligosaccharide-repeating units of the O-antigenic determinants was cloned and sequenced. A set of nucleotide primers(a forward and reverse) was selected to target a defined region of the guanosine diphospho-mannose(GDP-Man) pyrophosphorylase synthase gene : rfbM of Salmonella C serogroup. The primer set was used to develop a PCR-based rapid and specific detection system for Salmonella C1 serogroup. Amplification bands of predicted size(1,422bp) were generated from 11 different Salmonella C1 isolates. The bands were verified to be specific for the C1 serogroup by Southern blot analysis using reference homologous DNA specificity was further confirmed by the lack of reactivity with heterologous DNA derived from non-salmonella members of the family enterobacteriaeceae. A specificity of 100% was deduced along with a very high sensitivity shown by a detection limit of 1fg of a purified DNA template. The isolated DNA sequence was found to be 99.8% homologous to S montevideo but the related primers amplified with the predicted band sizes with all the Salmonella C1 serogroups tested. It is concluded that the PCR protocol based on the rfbM gene from S cholerasuis is optimal fast and specific for the detection of Salmonella C1 serogroup and also the corresponding probe is suitable for rapid detection of all Salmonella C1 serogroup DNA tested. This technology should facilitate the identification of contaminated pig products and for any other products contaminated with the Salmonalla C1 serogroup. The immediate impact of this developed method will be in the area of food safety of pig products with the potential prospect for adaptation to other food inspection technologies.

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A Utility-Based Hybrid Error Recovery Scheme for Multimedia Transmission over 3G Cellular Broadcast Networks (3G 방송망에서의 효율적인 멀티미디어 전송을 위한 유틸리티 기반 하이브라드 에러 복구기법)

  • Kang Kyung-Tae;Cho Yong-Jin;Cho Yong-Woo;Cho Jin-Sung;Shin Heon-Shik
    • Journal of KIISE:Information Networking
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    • v.33 no.4
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    • pp.333-342
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    • 2006
  • The cdma2000 lxEV - DO mobile communication system provides broadcast and multicast services (BCMCS) to meet an increasing demand from multimedia data services. The servicing of video streams over a BCMCS network must, however, face a challenge from the unreliable and error-prone nature of the radio channel. The BCMCS network uses Reed-Solomon coding integrated with the MAC protocol for error recovery. We analyze this coding technique and show that it is not effective in the case of slowly moving mobiles. To improve the playback quality of an MPEG-4 FGS video stream, we propose the Hybrid error recovery scheme, which combines Reed-Solomon with ARQ, using slots which are saved by reducing the Reed-Solomon coding overhead. The target packets to be retransmitted are prioritized by a utility function to reduce the packet error rate in the application layer within a fixed retransmission budget. This is achieved by considering of the map of the error control block at each mobile node. The proposed Hybrid error recovery scheme also uses the characteristics of MPEG-4 FGS (fine granularity scalability) to improve the video quality even when conditions are adverse: slow-moving nodes and a high error rate in the physical channel.

Interaction of Schwann Cells with Various Protein- or Polypeptide-Coated PLGA Surfaces (다양한 단백질과 폴리펩타이드로 코팅된 PLGA 표면과 슈반세포와의 상호관계)

  • Park Ki-Suk;Kim Su-Mi;Kim Moon-Suk;Lee Il-Woo;Rhee John-M.;Lee Hai-Bang;Khang Gil-Son
    • Polymer(Korea)
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    • v.30 no.5
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    • pp.445-452
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    • 2006
  • In this study, we investigated interaction of Schwann cells (SCs) with various cell-adhesive coated polymer surface. We used cell-adhesives that like a fibronectin (FN), fibrinogen(FG), laminin(LM), vitronectin (VN), poly-D-Iysine (PDL), and poly-L-Iysine (PLL) to coat PLGA film surface and evaluated the surface property of coated or not PLGA films by measurement of water contact angle and ESCA. SCs were cultured on coated or non-coated PLGA film surface, and then examined the cell adhesion and proliferation by cell count and SEM observation. Cell count results revealed initial cell adhesion related to protein adsorption on PLGA surface. In addition, serum content in media related to cell proliferation rate. In this result, we recognized that adhesion and proliferation of SCs were affected by specific cell-adhesives. In these results, we recognized that is important to provide the suitable surface environment according to cell types and culture condition for improvement of cell adhesion and proliferation.

Meat Species Identification using Loop-mediated Isothermal Amplification Assay Targeting Species-specific Mitochondrial DNA

  • Cho, Ae-Ri;Dong, Hee-Jin;Cho, Seongbeom
    • Food Science of Animal Resources
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    • v.34 no.6
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    • pp.799-807
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    • 2014
  • Meat source fraud and adulteration scandals have led to consumer demands for accurate meat identification methods. Nucleotide amplification assays have been proposed as an alternative method to protein-based assays for meat identification. In this study, we designed Loop-mediated isothermal amplification (LAMP) assays targeting species-specific mitochondrial DNA to identify and discriminate eight meat species; cattle, pig, horse, goat, sheep, chicken, duck, and turkey. The LAMP primer sets were designed and the target genes were discriminated according to their unique annealing temperature generated by annealing curve analysis. Their unique annealing temperatures were found to be $85.56{\pm}0.07^{\circ}C$ for cattle, $84.96{\pm}0.08^{\circ}C$ for pig, and $85.99{\pm}0.05^{\circ}C$ for horse in the BSE-LAMP set (Bos taurus, Sus scrofa domesticus and Equus caballus); $84.91{\pm}0.11^{\circ}C$ for goat and $83.90{\pm}0.11^{\circ}C$ for sheep in the CO-LAMP set (Capra hircus and Ovis aries); and $86.31{\pm}0.23^{\circ}C$ for chicken, $88.66{\pm}0.12^{\circ}C$ for duck, and $84.49{\pm}0.08^{\circ}C$ for turkey in the GAM-LAMP set (Gallus gallus, Anas platyrhynchos and Meleagris gallopavo). No cross-reactivity was observed in each set. The limits of detection (LODs) of the LAMP assays in raw and cooked meat were determined from $10pg/{\mu}L$ to $100fg/{\mu}L$ levels, and LODs in raw and cooked meat admixtures were determined from 0.01% to 0.0001% levels. The assays were performed within 30 min and showed greater sensitivity than that of the PCR assays. These novel LAMP assays provide a simple, rapid, accurate, and sensitive technology for discrimination of eight meat species.

Development of real-time PCR for rapid detection of Mycobacterium bovis DNA in cattle lymph nodes and differentiation of M. bovis and M. tuberculosis (소 림프절에서 Mycobacterium bovis DNA의 신속 검출과 M. bovis와 M. tuberculosis 감별을 위한 real-time PCR 개발)

  • Koh, Ba-Ra-Da;Jang, Young-Boo;Ku, Bok-Kyung;Cho, Ho-Seong;Bae, Seong-Yeol;Na, Ho-Myung;Park, Seong-Do;Kim, Yong-Hwan;Mun, Yong-Un
    • Korean Journal of Veterinary Service
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    • v.34 no.4
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    • pp.321-331
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    • 2011
  • Mycobacterium bovis, a member of the M. tuberculosis complex (MTC), is the causative agent of bovine tuberculosis. Detection of M. bovis and M. tuberculosis using conventional culture- and biochemical-based assays is time-consuming. Therefore, a simple and sensitive molecular assay for rapid detection would be of great help in specific situations such as faster diagnosis of bovine tuberculosis (bTB) infection in the abattoirs. We developed a novel multiplex real-time PCR assay which was applied directly to biological samples with evidence of bTB and it was allowed to differentiate between M. bovis and M. tuberculosis. The primers and TaqMan probes were designed to target the IS1081 gene, the multi-copy insertion element in the MTC and the 12.7-kb fragment which presents in M. tuberculosis, not in the M. bovis genome. The assay was optimized and validated by testing 10 species of mycobacteria including M. bovis and M. tuberculosis, and 10 other bacterial species such as Escherichia coli, and cattle lymph nodes (n=113). The tests identified 96.4% (27/28) as M. bovis from the MTC-positive bTB samples using conventional PCR for specific insertion elements IS1081. And MTC-negative bTB samples (n=85) were tested using conventional PCR and the real-time PCR. When comparative analyses were conducted on all bovine samples, using conventional PCR as the gold standard, the relative accuracy of real-time PCR was 99.1%, the relative specificity was 100%, and the agreement quotient (kappa) was 0.976. The detection limits of the real-time PCR assays for M. bovis and M. tuberculosis genomic DNA were 10 fg and 0.1 pg per PCR reaction, respectively. Consequently, this multiplex real-time PCR assay is a useful diagnotic tool for the identification of MTC and differentiation of M. bovis and M. tuberculosis, as well as the epidemiologic surveillance of animals slaughtered in abattoir.

Studies on the Regional Distribution and Some Ecological Characteristics of Chinese White-wax Scale in Korea (국내에서 서식하는 쥐똥밀깍지벌레(Ericerus pela)의 분포 및 생태에 관한 연구)

  • 박인규;마영일;윤형주;양성열
    • Korean journal of applied entomology
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    • v.37 no.2
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    • pp.137-142
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    • 1998
  • The regional distribution and some ecological characteristics of Chinese white-wax scale, Ericerus pela Chavannes, were investigated from 1996 to 1997. It was found that 10.8% of the stems of the privet, Ligustrum obtusifolium infested with the scale in Chtingpyijng, whereas only 1.1% of them infested in Y6ngwol and Pyongtaek. The average survival rate of the female adult marked 85.3% after it hibernated on the privet, Ligustrum obtusifolium. A female laid 7,783.5 eggs in average and 36.7% of females fell on the range of 7,000-10,000 eggs. It sized 0.40 mm in length and 0.21 mm in width. The hatchability was highest at 27$^{\circ}$C with 66.8% and it seemed the optimum temperature for incubation. The pupation rate was lower than 50.0% at the above experimental temperatures and the emergence rate marked 67.3% at 25$^{\circ}$C. When the egg was preserved at the various low temperatures, it was found that the egg could be preserved at lS$^{\circ}$C for 50 days in maximum.

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Effects of Kimchi Supplementation and/or Exercise Training on Body Composition and Plasma Lipids in Obese Middle School Girls (운동 및 김치 보충제 섭취가 비만 여중생의 신체조성 및 혈중지질에 미치는 영향)

  • 백영호;곽정록;김세종;한성섭;송영옥
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.30 no.5
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    • pp.906-912
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    • 2001
  • The effects of aerobic exercises and/or supplementation of kimchi on changes of the body composition and plasma lipids of obese middle school girls were studied. Thirty eight girls, 28 obese girls and 10 normal weighed girls, were paricipated. Among obese girls, 8 were assigned to exercise group (FG) 12 were grouped as kimchi group (KG) , and 8 were asked to practice excercise and to take kimchi simultaneously(excercise kimchi group, EKG). Ten girls whose weight is normal asked to remain o their own diet during 6 weeks of experiment (control group, CG)/ EG practiced jogging and rope-jumping for 60 minutes four times a week and KG took 3 g of freeze-dried kimchi packed in a 500 mg capsule daily which is equivalent to 30 g of fresh kimchi, EKC, EG and KG showed beneficial effects on changes of the body composition and plasma lipids compared to those of CG, EG showed greater effect than KG in reducing body fat resulted decrease in BMI, fat mas,. abdominal fat, and triglyceride concentration and increase in HDL-cholesterol. KG seemed to have greater effect on lowering plasma cholesterol and LDL-cholesterol than EG/ But the greatest effects in terms of reduction in weight, BMI fat mass, abdominal fat, total cholesterol, LDL-cholesterol, triglyceride, and increase in HDL-cholesterol were observed from EKG. These results indicate that kimchi supplemenation while practicing excercise might improve the obese state by reducing body fat content as well as reducing plasma lipids.

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Analysis of the Genome Sequence of Strain GiC-126 of Gloeostereum incarnatum with Genetic Linkage Map

  • Jiang, Wan-Zhu;Yao, Fang-Jie;Fang, Ming;Lu, Li-Xin;Zhang, You-Min;Wang, Peng;Meng, Jing-Jing;Lu, Jia;Ma, Xiao-Xu;He, Qi;Shao, Kai-Sheng;Khan, Asif Ali;Wei, Yun-Hui
    • Mycobiology
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    • v.49 no.4
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    • pp.406-420
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    • 2021
  • Gloeostereum incarnatum has edible and medicinal value and was first cultivated and domesticated in China. We sequenced the G. incarnatum monokaryotic strain GiC-126 on an Illumina HiSeq X Ten system and obtained a 34.52-Mb genome assembly sequence that encoded 16,895 predicted genes. We combined the GiC-126 genome with the published genome of G. incarnatum strain CCMJ2665 to construct a genetic linkage map (GiC-126 genome) that had 10 linkage groups (LGs), and the 15 assembly sequences of CCMJ2665 were integrated into 8 LGs. We identified 1912 simple sequence repeat (SSR) loci and detected 700 genes containing 768 SSRs in the genome; 65 and 100 of them were annotated with gene ontology (GO) terms and KEGG pathways, respectively. Carbohydrate-active enzymes (CAZymes) were identified in 20 fungal genomes and annotated; among them, 144 CAZymes were annotated in the GiC-126 genome. The A mating-type locus (MAT-A) of G. incarnatum was located on scaffold885 at 38.9 cM of LG1 and was flanked by two homeodomain (HD1) genes, mip and beta-fg. Fourteen segregation distortion markers were detected in the genetic linkage map, all of which were skewed toward the parent GiC-126. They formed three segregation distortion regions (SDR1-SDR3), and 22 predictive genes were found in scaffold1920 where three segregation distortion markers were located in SDR1. In this study, we corrected and updated the genomic information of G. incarnatum. Our results will provide a theoretical basis for fine gene mapping, functional gene cloning, and genetic breeding the follow-up of G. incarnatum.