• Title/Summary/Keyword: One-step

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A Combination Strategy for Construction of Peptide-β2m-H-2Kb Single Chain with Overlap Extension PCR and One-Step Cloning

  • Xu, Tao;Li, Xiaoe;Wu, You;Shahzad, Khawar Ali;Wang, Wei;Zhang, Lei;Shen, Chuanlai
    • Journal of Microbiology and Biotechnology
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    • v.26 no.12
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    • pp.2184-2191
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    • 2016
  • The time-consuming and high-cost preparation of soluble peptide-major histocompatibility complexes (pMHC) currently limits their wide uses in monitoring antigen-specific T cells. The single-chain trimer (SCT) of peptide-${\beta}2m$-MHC class I heavy chain was developed as an alternative strategy, but its gene fusion is hindered in many cases owing to the incompatibility between the multiple restriction enzymes and the restriction endonuclease sites of plasmid vectors. In this study, overlap extension PCR and one-step cloning were adopted to overcome this restriction. The SCT gene of the $OVA_{257-264}$ peptide-$(GS_4)_3-{\beta}2m-(GS_4)_4-H-2K^b$ heavy chain was constructed and inserted into plasmid pET28a by overlap extension PCR and one-step cloning, without the requirement of restriction enzymes. The SCT protein was expressed in Escherichia coli, and then purified and refolded. The resulting $H-2K^b/OVA_{257-264}$ complex showed the correct structural conformation and capability to bind with $OVA_{257-264}$-specific T-cell receptor. The overlap extension PCR and one-step cloning ensure the construction of single-chain MHC class I molecules associated with random epitopes, and will facilitate the preparation of soluble pMHC multimers.

Application of a nested-polymerase chain reaction assay to differentiate the genotypes of porcine circovirus 2 (돼지 써코바이러스 2형 유전형 분류를 위한 nested-PCR 적용)

  • Chu, Keum-Suk
    • Korean Journal of Veterinary Service
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    • v.34 no.1
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    • pp.13-18
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    • 2011
  • The purpose of this study was to apply a nested-polymerase chain reaction (nPCR) assay to detect and differentiate PCV 2a and PCV 2b. The compared with nPCR and one-step PCR and nPCR showed more sensitive in the detection of PCV-2 from tissue and blood samples. The total of 52 tissue samples was collected from postweanning pigs from 2006 to 2010. All tissue samples showed positive for PCV-2 in one-step PCR and nPCR, followed by the nPCR in order to identify the genotypes of PCV-2. 2 samples (3.8%) showed positive for PCV 2a, and 35 samples were positive for PCV 2b (67.3%), 15 samples (28.9%) were positive the dual genotypes. In addition, 42 blood samples which were collected from the 5 different swine farms were compared figure out the detection rates of nPCR and one-step PCR. The PCV 2 was positive by one-step PCR in 21 samples (50.0%) and nPCR was positive in 37 samples (88.1%). The PCV 2 genotypes in blood samples and 32 samples (76.2%) were positive for PCV 2b and none were positive for PCV 2a, 5 samples (11.9%) were positive for dual genotypes. These results suggest that the nPCR is very efficient for genotyping blood samples and differentiating the genotypes of PCV-2 from field samples.

Development of a One-Step Duplex RT-PCR Method for the Simultaneous Detection of VP3/VP1 and VP1/P2B Regions of the Hepatitis A Virus

  • Kim, Mi-Ju;Lee, Shin-Young;Kim, Hyun-Joong;Lee, Jeong Su;Joo, In Sun;Kwak, Hyo Sun;Kim, Hae-Yeong
    • Journal of Microbiology and Biotechnology
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    • v.26 no.8
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    • pp.1398-1403
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    • 2016
  • The simultaneous detection and accurate identification of hepatitis A virus (HAV) is critical in food safety and epidemiological studies to prevent the spread of HAV outbreaks. Towards this goal, a one-step duplex reverse-transcription (RT)-PCR method was developed targeting the VP1/P2B and VP3/VP1 regions of the HAV genome for the qualitative detection of HAV. An HAV RT-qPCR standard curve was produced for the quantification of HAV RNA. The detection limit of the duplex RT-PCR method was 2.8 × 101 copies of HAV. The PCR products enabled HAV genotyping analysis through DNA sequencing, which can be applied for epidemiological investigations. The ability of this duplex RT-PCR method to detect HAV was evaluated with HAV-spiked samples of fresh lettuce, frozen strawberries, and oysters. The limit of detection of the one-step duplex RT-PCR for each food model was 9.4 × 102 copies/20 g fresh lettuce, 9.7 × 103 copies/20 g frozen strawberries, and 4.1 × 103 copies/1.5 g oysters. Use of a one-step duplex RT-PCR method has advantages such as shorter time, decreased cost, and decreased labor owing to the single amplification reaction instead of four amplifications necessary for nested RT-PCR.

Fixing Behaviors of Dimethylamino Anthraquinone Disperse Dyes and Monochlorotriazinyl Azo Reactive Dyes on P/C Blended Fabrics in One-Step Printing (디메틸아미노안트라퀴논계 분산염료와 아조계 모노클로로트리아진형 반응염료에 의한 P/C혼방직물의 일단계 날염에 있어 고착거동)

  • Park, Geon-Yong;Seo, Gi-Sung
    • Textile Coloration and Finishing
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    • v.19 no.3
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    • pp.18-25
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    • 2007
  • The fixing behaviors of anthraquinone disperse dyes containing dimethylamino substituent, such as C. I. Disperse Violet 26(D.V.26) and C. I. Disperse Blue 14(D.V.14), or containing diamino substituent, such as C. I. Disperse Blue 73(D.B.73), and monochlorotriazinyl azo reactive dyes, such as C. I. Reactive Orange 13(R.O.13), C. I. Reactive Red 3(R.R.3). C. I. Reactive Yellow 2(R.Y.2) on polyester/cotton blend(P/C) fabrics were examined for the one-step printing of P/C fabrics. The high temperature steaming of $175^{\circ}C$ is the most satisfactory fixing method for P/C one-step printing with above disperse and reactive dyes among the four different fixing methods: $175^{\circ}C$ steaming, $102^{\circ}C$ steaming${\rightarrow}175^{\circ}C$ steaming, $190^{\circ}C$ thermosol, $102^{\circ}C$ steaming${\rightarrow}190^{\circ}C$ thermosol. $190^{\circ}C$ thermosol is unfit to fix R.R.3 and R.Y.2 whose heat stability is poor. It was found that D.V.26 and D.B.14 containing dimethylamino substituent are unstable for heat and alkali, but D.B.73 is stable for them to print P/C blend fabrics with R.O.13 which is also stable for heat. Therefore we found that D.B.73, R.O.13 and a pair of D.B.73 and R.O.13 were very suitable for one-step printing of P/C blend fabrics.

Effect of additional etching and ethanol-wet bonding on the dentin bond strength of one-step self-etch adhesives

  • Ahn, Joonghee;Jung, Kyoung-Hwa;Son, Sung-Ae;Hur, Bock;Kwon, Yong-Hoon;Park, Jeong-Kil
    • Restorative Dentistry and Endodontics
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    • v.40 no.1
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    • pp.68-74
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    • 2015
  • Objectives: This study examined the effects of additional acid etching on the dentin bond strength of one-step self-etch adhesives with different compositions and pH. The effect of ethanol wetting on etched dentin bond strength of self-etch adhesives was also evaluated. Materials and Methods: Forty-two human permanent molars were classified into 21 groups according to the adhesive types (Clearfil SE Bond [SE, control]; G-aenial Bond [GB]; Xeno V [XV]; Beauti Bond [BB]; Adper Easy Bond [AE]; Single Bond Universal [SU]; All Bond Universal [AU]), and the dentin conditioning methods. Composite resins were placed on the dentin surfaces, and the teeth were sectioned. The microtensile bond strength was measured, and the failure mode of the fractured specimens was examined. The data were analyzed statistically using two-way ANOVA and Duncan's post hoc test. Results: In GB, XV and SE ($pH{\leq}2$), the bond strength was decreased significantly when the dentin was etched (p < 0.05). In BB, AE and SU (pH 2.4 - 2.7), additional etching did not affect the bond strength (p > 0.05). In AU (pH = 3.2), additional etching increased the bond strength significantly (p < 0.05). When adhesives were applied to the acid etched dentin with ethanol-wet bonding, the bond strength was significantly higher than that of the no ethanol-wet bonding groups, and the incidence of cohesive failure was increased. Conclusions: The effect of additional acid etching on the dentin bond strength was influenced by the pH of one-step self-etch adhesives. Ethanol wetting on etched dentin could create a stronger bonding performance of one-step self-etch adhesives for acid etched dentin.

On the Equivalance of Some Fixed Point Iterations

  • Ozdemir, Murat;Akbulut, Sezgin
    • Kyungpook Mathematical Journal
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    • v.46 no.2
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    • pp.211-217
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    • 2006
  • In this paper, we have shown that the convergence of one-step, two-step and three-step iterations is equivalent, which are known as Mann, Ishikawa and Noor iteration procedures, for a special class of Lipschitzian operators defined in a closed, convex subset of an arbitrary Banach space.

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COLOR STABILITY AND SURFACE TEXTURE TO TEMPERATURE AT STASHING OF PORCELAIN RESTORATION (도재수복물의 표면첨색시 온도에 따른 색조안정성과 표면조도의 평가)

  • Beak Eun-Ju;Bae Tae-Seong;Song Kwang-Yeob;Park Charn-Woon
    • The Journal of Korean Academy of Prosthodontics
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    • v.29 no.3
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    • pp.87-99
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    • 1991
  • The effect of oven firing on the color stability and surface texture of extrinsic stains used characterization and color modification of metal cermic restoration were studied by comparing different temperatures and techniques. They were autoglaze technique, applied glaze technique, two step low-fusing glaze technique and one step low-fusing glaze technique. Autoglaze technique and applied glaze technique were air-fired to manufacturer's recommended temperature. Two step low-fusing glaze technique is separated staining and overglazing. Thin slurry of stain powder were air-fired to $1500^{\circ}F$, Overglaze powder powder was then mixed with seam medium and air-fired to $170^{\circ}F$. One step low-fusing glaze technique is combined staining and overglazing. Thin slurry of stain and glaze powder were air-fired to $170^{\circ}F$. The obtained results were as fellows. 1. Slightly significant color differences by colorimeter were found between different stain application and fusion techniques(P<0.05). 2. Two step low-fusing glaze method showed the most rough surface, especially ceramco orage stain(P<0.05). 3. Surface roughness increased gradually in order of autoglaze, applied glaze, one step low-fusing glaze but they were not significantly different(P<0.05). 4. When two step low-fusing glaze was applied, both color measurement and surface texture were sinificantly different from other groups(P<0.05).

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Analysis of Two-step programming characteristics of the flash EEPROM's (Flash EEPROM의 two-step 프로그램 특성 분석)

  • 이재호;김병일;박근형;김남수;이형규
    • Journal of the Korean Institute of Telematics and Electronics D
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    • v.34D no.9
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    • pp.56-63
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    • 1997
  • There generally exists a large variation in the thereshold voltages of the flash EEPROM cells after they are erased by using th fowler-nordheim tunneling, thereby getting some cells to be overeased. If the overerased cells are programmed with the conventional one-step programming scheme where an 12-13V pulse with the duration of 100.mu.S is applie don the control gate for the programming, they can suffer from the significant degradation of the reliability of the gate oxide. A two-step programming schem, where an 8/12 V pulse with a duration of 50.mu.S for each voltage is applied on the control gate for the programming, has been studied to solve the problem. The experimental results hav eshown that there is little difference in the programming characteristics between those two schemes, whereas the degradation of the gate oxide due to the programming can be significantly reduced with the two-step programming scheme compared to that with the one-step programming scheme. This is possibly because the positive charge stored in the floating gate of the overerased cells is compensate dwith the electrons injected into the floating gate while the 8V pulse is applied on the control gate, which leaves the overerased cells in the normally erased state after the duration of the 8V pulse.

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A Study on Internship Program Development for Fashion Industry - Focused on Internship Activation Method of Fashion Industry and Government- (패션산업 인턴십 프로그램 개발에 관한 연구 - 패션기업과 정부의 인턴십 활성화 방안을 중심으로 -)

  • Yu, Ji-Hun;Chung, Sang-Gil
    • The Research Journal of the Costume Culture
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    • v.13 no.5 s.58
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    • pp.699-711
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    • 2005
  • This study was following one of 'A study on the consciousness of fashion industries internship'. The purposes of this study were to develop the internship program which focused on practical use to introduce and carry out for fashion industries, and secondly to propose some regime for government to activate fashion internship. Reference searching method and depth interviewing method were used for this study. The results were as follows : Fashion industry internship was grouped into two classes, 'on-the-job training'; educating students fields and 'talent hunting'; selecting good persons. Internship of industry-academic world was classified into two types; the one is 'credit type' which has curriculums between universities and industries and the other is 'non-credit type' which has not any credit and is operated by industry own system. This study provided the development courses of pragmatic program to perform internship systematically and it also provided the program models for guide line in fashion industries. Six grades such as ready step, introduction step, selection step, management step, evaluation step and feed-back step were proposed for the internship program development steps of fashion industries. A virtual organization, 'The Fashion Industry and Academy Association' was proposed as a policy for activating internship between universities, industries and government.

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Concentration Distributions and A Reduction Strategy of Airborne Radon in Seoul Metropolitan Subway Stations (서울시 지하철역내의 라돈 농도분포 및 저감대책)

  • 김동술;김윤신;김신도;신응배;김성천;유정석
    • Journal of Korean Society for Atmospheric Environment
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    • v.9 no.4
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    • pp.271-277
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    • 1993
  • Indoor radon has been known as one of the notorious carcinogens. However, a safe environmental criterion of radon has not yet been established in Korea, The main objectives of this study were to study concentration distributions of radon, to trace radon sources in subways, and to obtain a strategy for radon reduction in Seoul metropolitan area. Radon concentrations had been extensively determined by several steps. The first step was to survey radon levels in all of 83 subway stations from October to November in 1991. The second step was to select 40 out of 83 stations and then to study seasonal variations in 1991 and 1992. The third step was to monitor radon levels by hourly-basis plans. The fourth step was to seek a radon reduction strategy by altering ventilation at Ankuk station where had the highest radon concentration during the first measurement step. Each underground floor in the station was divided into 10 sites to measure hourly radon variations. The final step of the study was to measure radon concentrations in groundwater that is one of the possible main sources radon place. The result of the various measuring approaches showed short-and long-term radon variation and indicated radon reduction schemes.

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