• Title/Summary/Keyword: primer

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Genetic analysis of norovirueses in Busan (부산지역 노로바이러스의 유전적 분석)

  • Kim, Kwang-Il;Jin, Ji-Woong;Jeong, Hyun-Do
    • Journal of fish pathology
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    • v.24 no.3
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    • pp.255-268
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    • 2011
  • For detection of noroviruses (NVs), we compared various PCR primer sets based on reverse transcription nested PCR (RT-nested PCR) in the water samples from Dong brook in Busan, South Korea. We designed various new primer sets based on the most conserved sequences of the capsid protein gene that react with diverse NVs found in Korea. Designed primer sets (KG1F/KG1R and KG2F/KG2R, named as PNK) for the respective genogroups of NVs, genogroup I and II (GI and GII), were applied to detect NVs in the water samples from Dong brook concentrated with ultracentrifugation. In the application to the water samples, proportion of GI (76.47%) and GII (70.59%) in water samples of Dong brook in RT-nested PCR with the primer sets of this study. However, no significant differences of the proportion of the positive samples were not found between RT-nested PCRs with reported and newly designed primer sets. From the nucleotide sequencing, GI and GII of NVs present in Dong brook were appeared to be the members of 1/2/4/5/9/10 genotypes, and 3/4/5/11/13 genotypes respectively. Appeared genotype 4 of GII known as an one of main genotype found in patients of many Asian countries warned us to consider the risks of norovirus in aquatic environments in southern part of Korea.

EFFECT OF THERMOCYCLING ON BONDING OF COMPOSITE RESIN AFTER DENTIN SURFACE TREATMENTS (상아질 표면처리후 복합레진 수복시 온도변화가 결합에 미치는 영향)

  • Lee, Yong-Woo;Son, Ho-Hyun
    • Restorative Dentistry and Endodontics
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    • v.16 no.2
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    • pp.126-142
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    • 1991
  • The purpose of this study was to evaluate the effect of various dentin surface treatments on shear bond strength, microhardness and fracture mode before and after thermocycling. Recently extracted 75 human molars were used. The teeth were sagittal sectioned faciolingually to obtain 150 specimens. They were randomly divided into six groups. Mesial and distal dentinal surfaces of specimens were exposed by grinding and treated respectively with GC-DENTIN CONDITIONER. 10-3 solution of 4-Meta, Cleansar and Primer of GLUMA, Scotchprep of Scotchbond 2, DENTIN CONDITIONER and PRIMER A, B of ALL BOND according to the manufacturers directions. Specimens of one group were not treated. Adhesive agent of Scotchbond 2, were applied and cured on the treated dentin surfaces. After P-50 were cured on them, specimens were stored in 31c water for 24 hours before shear bond strength measurement Shear bond strength was measured in 10 specimens of each group. 10 specimens of each group were thermocycled in $20^{\circ}C$, $60^{\circ}C$,$20^{\circ}C$, $4^{\circ}C$, $20^{\circ}C$ water in order, for 30 seconds respectively, 100 times a day for 7 days. After thermocycling shear bond strength was measured. Microhardness was checked on treated dentin surface and fractured dentin surface in 10 specimens respectievly. Francture modes were observed with SEM The following results were obtained. 1. Before thermocycling. shear bond strengths in the specimens treated with DENTIN CONDITIONER and PRIMER A, B of ALL BOND were significantly higher than those in other specimens(P<0.01). 2. After thermocycling. shear bond strengths in the specimens treated with Cleanser and Primer of GLUMA, Scotchprep of Scotchbond 2 and DENTIN CONDITIONER and PRIMER A, B of AIL BOND were significantly higher than those in specimens not: treated, treated with GC-DENTIN CONDITIONER and 10-3 solution of 4-Meta(P<0.01). Shear bond strengths in the specimens treated with GC-DENTIN CONDITIONER and PRIMER A, B of ALL BOND were significantly higher than those in other specimens except those treated with Scotchprep of Srotchbond 2(P<0.01). 3. Shear bond strengths after thermocycling were reduced in the specimens not treated, treated with GC-DENTIN CONDITIONER and 10-3 solution of 4-Meta and were increased in the specimens treated with Cleanser and Primer of GLUMA, Scotchprep of Scotchbond 2, without significance, compared with those before thermocycling. In the specimens treated with DENTIN CONDITIONER and PRIMER A, B of ALL BOND, shear bond strengths after thermocycling were significantly increased, compared with those before thermocycling(P<0.01). 4. Microhardnesses in the fractured surfaces after shear bond strength measurement were significantly increased in the specimens treated with 10-3 solution of 4-Meta and significantly decreased in the specimens treated with DENTIN CONDITIONER and PRIMER A, B of ALL BOND, compared with those in the treated dentin surfaces(P<0.01). 5. In the specimens treated with Cleanser and Primer of GLUMA, Scotchprep of Scotchbond 2 and DENTIN CONDITIONER and PRIMER A, B of ALL BOND, cohesive fracture modes were observed more than adhesive fracture modes.

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Detection of Nocardia sp. Hl7-1 by PCR during Bioremediation of Crude Oil-Contaminated Soil (원유 오염토양의 Bioremediation과정 동안 PCR을 이용한 Nocardia sp. Hl7-1의 검출)

  • Baek, Kyung-Hwa;Lee, Young-Ki;Lee, In-Sook;Oh, Hee-Mock;Yoon, Byung-Dae;Kim, Hee-Sik
    • Microbiology and Biotechnology Letters
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    • v.32 no.1
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    • pp.91-95
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    • 2004
  • For the detection of the oil-degrading bacterium, Nocardia sp. Hl7-1, inoculated during the bioremediation of oil-contaminated soil, a species-specific primer was constructed based on the 16S rDNA sequence of this strain. Two forward primers and two reverse primers were designed and tested against both closely and distantly related bacterial strains. All the primers designed were specific to the Nocardia sp. H17-1. Particularly, primer sets NH169F-NH972R and NH575F-NH972R could be used to detect 50 fg of template DNA and TEX>$1.2${\times}$10^4$ CFU/g of sandy soil. These two PCR primer sets successfully detected the H 17-1 strain in the oil-con-laminated soil samples containing heterogeneous DNA. We also conformed the primer specificity by restriction-enzyme cleavage of the PCR products and denaturing gradient gel electrophoresis.

EFFECT OF APPLICATION METHODS OF A SELF-ETCHING PRIMER ADHESIVE SYSTEM ON ENAMEL BOND STRENGTH (자가부식 프라이머 접착제의 적용방식이 법랑질의 결합강도에 미치는 영향)

  • Park, Jae-Gu;Cho, Kwon-Hwan;Cho, Young-Gon
    • Restorative Dentistry and Endodontics
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    • v.33 no.2
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    • pp.90-97
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    • 2008
  • The purpose of this study was to evaluate the effect of passive or active application of primer and coat times of bond on the shear bond strength when a self-etching primer adhesive (Clearfil SE Bond) was applied to enamel surface. Crowns of sixteen human molars were selected. Buccal and lingual enamels of crowns were partially exposed and slabs of 1.2 mm thick were made. They were divided into one of four equal groups (n = 8). Group 1: passive application of Primer and 1 coat of Bond, Group 2: active application of Primer and 1 coat of Bond, Group 3: passive application of Primer and 2 coats of Bond, Group 4: active application of Primer and 2 coats of Bond. Clearfil AP-X was bonded to enamel suface of each group using Tygon tubes. The bonded specimens were subjected to microshear bond strength (uSBS) testing with a crosshead speed of 1 mm/min. The results of this study were as follows; 1. The uSBS of Group 1 was the lowest among groups and the uSBS of Group 4 was the highest. 2. There was not statistically significant interaction between enamel uSBS by application method of Primer and coat time of Bond (p > 0.05). 3. There was not statistically significant difference between enamel uSBS by passive and active application of Primer (p > 0.05). 4. There was statistically significant difference between enamel uSBS by one- and two-coat of Bond (p < 0.05).

Development of a psychrophilic-SCAR marker for Pleurotus eryngii (큰느타리버섯의 저온적응성 형질에 관련된 SCAR Marker 개발)

  • Kim, Su Chul;Hwang, Hye Sung;Cho, Yun Jun;Kim, Hye Su;Ryu, Jae-San;Cho, Soo Jeong
    • Journal of Mushroom
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    • v.11 no.3
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    • pp.171-176
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    • 2013
  • Genomic DNAs of psychrophilic strains of Pleurotus eryngii were analyzed by randomly amplified polymorphic DNA (RAPD) using OP-A, OP-B, OP-L, OP-P, OP-R and OP-S3 primers to develop the strain-specific DNA marker. A unique DNA fragment with the size of 480 bp was yielded by OP-S3 primer from the psychrophilic strain. A sequence characterized amplified region (SCAR) marker, designated as OP-S3-1, was designed on the basis of the determined sequence. The PCR analysis with the OP-S3-1 primer showed that this SCAR marker can clearly distinguish the psychrophilic strains from the control strains.

Species-specific Marker of Phytophthora pinifolia for Plant Quarantine in Korea (국내 식물검역대상 Phytophthora pinifolia의 PCR 검출을 위한 종 특이적 마커 개발)

  • Kim, Narae;Choi, You Ri;Seo, Mun Won;Song, Jeong Young;Kim, Hong Gi
    • The Korean Journal of Mycology
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    • v.44 no.2
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    • pp.103-107
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    • 2016
  • To establish a rapid and accurate detection of Phytophthora pinifolia, which is a quarantine pathogenic fungus in Korea, a species-specific primer was developed based on the ras-related protein (Ypt1) gene. Species-specific primer based on the DNA sequences of Ypt1 gene amplified 193 bp polymerase chain reaction (PCR) product for P. pinifolia. The primer pair yielded the predicted PCR product size exactly in testing with target pathogen DNAs, but not from the other 10 species of Phytophthora and 14 species of other phytopathogenic fungi. The primer pair also showed only the species-specific amplification curve on realtime PCR on target pathogen DNA. The detection sensitivity of real time PCR using species-specific primer pair was 10 to 100 times higher than conventional PCR, with 1 to $10pg/{\mu}L$.

Application of LATE-PCR to Detect Candida and Aspergillus Fungal Pathogens by a DNA Hybridization Assay

  • Gopal, Dhayaalini Bala;Lim, Chua Ang;Khaithir, Tzar Mohd Nizam;Santhanam, Jacinta
    • Microbiology and Biotechnology Letters
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    • v.45 no.4
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    • pp.358-364
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    • 2017
  • Asymmetric PCR preferentially amplifies one DNA strand for use in DNA hybridization studies. Linear-After-The-Exponential-PCR (LATE-PCR) is an advanced asymmetric PCR method which uses innovatively designed primers at different concentrations. This study aimed to optimise LATE-PCR parameters to produce single-stranded DNA of Candida spp. and Aspergillus spp. for detection via probe hybridisation. The internal transcribed spacer (ITS) region was used to design limiting primer and excess primer for LATE-PCR. Primer annealing and melting temperature, difference of melting temperature between limiting and excess primer and concentration of primers were optimized. In order to confirm the presence of single-stranded DNA, the LATE-PCR product was hybridised with digoxigenin labeled complementary oligonucleotide probe specific for each fungal genus and detected using anti-digoxigenin antibody by dot blotting. Important parameters that determine the production of single-stranded DNA in a LATE-PCR reaction are difference of melting temperature between the limiting and excess primer of at least $5^{\circ}C$ and primer concentration ratio of excess primer to limiting primer at 20:1. LATE-PCR products of Candida albicans, Candida parapsilosis, Candida tropicalis and Aspergillus terreus at up to 1:100 dilution and after 1 h hybridization time, successfully hybridised to respective oligonucleotide probes with no cross reactivity observed between each fungal genus probe and non-target products. For Aspergillus fumigatus, LATE-PCR products were detected at 1:10 dilution and after overnight hybridisation. These results indicate high detection sensitivity for single-stranded DNA produced by LATE-PCR. In conclusion, this advancement of PCR may be utilised to detect fungal pathogens which can aid the diagnosis of invasive fungal disease.

Microtensile bond strength of resin cement primer containing nanoparticles of silver (NAg) and amorphous calcium phosphate (NACP) to human dentin

  • Arjmand, Nushin;Boruziniat, Alireza;Zakeri, Majid;Mohammadipour, Hamideh Sadat
    • The Journal of Advanced Prosthodontics
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    • v.10 no.3
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    • pp.177-183
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    • 2018
  • PURPOSE. The purpose of the current study was to evaluate the effect of incorporating nanoparticles of silver (NAg) and amorphous calcium phosphate (NACP) into a self-etching primer of a resin cement on the microtensile bond strength of dentin, regarding the proven antibacterial feature of NAg and remineralizing effect of NACP. MATERIALS AND METHODS. Flat, mid-coronal dentin from 20 intact extracted human third molars were prepared for cementation using Panavia F2.0 cement. The teeth were randomly divided into the four test groups (n=5) according to the experimental cement primer composition: cement primer without change (control group), primer with 1% (wt) of NACP, primer with 1% (wt) of physical mixture of NACP+Nag, and primer with 1% (wt) of chemical mixture of NACP+Nag. The resin cement was used according to the manufacturer's instructions. After storage in distilled water at $37^{\circ}C$ for 24 h, the bonded samples were sectioned longitudinally to produce $1.0{\times}1.0mm$ beams for micro-tensile bond strength testing in a universal testing machine. Failure modes at the dentin-resin interface were observed using a stereomicroscope. The data were analyzed by one-way ANOVA and Tukey's post-hoc tests and the level of significance was set at 0.05. RESULTS. The lowest mean microtensile bond strength was obtained for the NACP group. Tukey's test showed that the bond strength of the control group was significantly higher than those of the other experimental groups, except for group 4 (chemical mixture of NACP and NAg; P=.67). CONCLUSION. Novel chemical incorporation of NAg-NACP into the self-etching primer of resin cement does not compromise the dentin bond strength.

EFFECT OF METAL PRIMER TREATMENT OF THE Au-Ag-PD ALLOY SURFACE ON THE METAL-RESIN BONDING (치과용 금-은-팔라디움 합금에 대한 프라이머 처리가 금속-레진 접착에 미치는 영향)

  • Lee Kang;Lee Cheong-Hee;Jo Kwang-Hun;Kim Kyo-Han
    • The Journal of Korean Academy of Prosthodontics
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    • v.39 no.4
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    • pp.417-432
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    • 2001
  • The pcf metal primers on the bond strength and durability of 4-META/MMA-TBB resins adhered to an Au-Ag-Pd alloy. For this study, the specimens were divided into 8 groups as follows: Thermocyle 0 : (1) control group : sandblast, (2) Group I : sandblast + Cesead Opaque Primer; (3) Group II : sandblast + Metal Primer; (4) Group III : sandblast + V-Primer; Thermocyle 10,000 (5) control sandblast: (6) Group I : sandblast + Cesead Opaque Primer: (7) Group II : sandblast + Metal Primer; (8) Group III sandblast + V-Primer. The shear bond strength was determined using an Instron were observed with the use of scanning electron microscope. Finally, the strengths of bonded joints were evaluated with regard to their adherence energy using a wedge test. The results obtained were as follows ; (1) The shear bond strength of 4-META/MMA-TBB resin to the Au-Ag-Pd alloy was significantly improved in all the groups treated with the primers (p<0.05). (2) Regardless of the adhesive primers used, a significant difference was observed in the bond strength of the thermocycle 0 groups and 10,000 groups (p<0.05). (3) Both before and after thermocycling, the strongest bond strength between the resin an the alloy was obtained after treatment with a metal primer containing MEPS (p<0.05). (4) In the wedge test, the adherence energies of the control group and Group III decreased more rapidly than those of Group I and II during the 2nd day of storage in water.

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Highly Specific Detection of Five Exotic Quarantine Plant Viruses using RT-PCR

  • Choi, Hoseong;Cho, Won Kyong;Yu, Jisuk;Lee, Jong-Seung;Kim, Kook-Hyung
    • The Plant Pathology Journal
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    • v.29 no.1
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    • pp.99-104
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    • 2013
  • To detect five plant viruses (Beet black scorch virus, Beet necrotic yellow vein virus, Eggplant mottled dwarf virus, Pelargonium zonate spot virus, and Rice yellow mottle virus) for quarantine purposes, we designed 15 RT-PCR primer sets. Primer design was based on the nucleotide sequence of the coat protein gene, which is highly conserved within species. All but one primer set successfully amplified the targets, and gradient PCRs indicated that the optimal temperature for the 14 useful primer sets was $51.9^{\circ}C$. Some primer sets worked well regardless of annealing temperature while others required a very specific annealing temperature. A primer specificity test using plant total RNAs and cDNAs of other plant virus-infected samples demonstrated that the designed primer sets were highly specific and generated reproducible results. The newly developed RT-PCR primer sets would be useful for quarantine inspections aimed at preventing the entry of exotic plant viruses into Korea.