• Tuberculosis and Respiratory Diseases
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• 제39권6호
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• pp.517-525
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• 1992

연구배경 : 1985년 Saiki등에 의해, 특정한 DNA를 연속적으로 복제할 수 있는 방법인 polymerase chain reaction (PCR)이 개발된 이래, PCR은 검체내에 극미량으로 존재하고 있는 병원체의 진단에 큰 도움을 줄 것으로 기대되었다. 결핵균의 진단방법중, 도말염색 방법은 감수성이 낮아서 문제가 되고 있으며, 배양은 감수성은 높으나 기간이 오래 걸려서 임상적으로 도움을 주지 못하는 경우가 많다. 이에 저자들은 Mycobacterium tuberculosis의 특이 단백질인 65 kD mycobacterial antigen을 encoding하는 2520 base pair DNA중, 383 base pair DNA를 이용한 PCR과 IS6110 fragment의 일부인 123 base pair DNA를 이용한 PCR을 시행하여, 이의 감수성과 특이도를 알아보고 폐결핵 환자의 객담을 검체로한 결핵의 조기진단 방법을 개발하고자 하였다. 방법 : M. tuberculosis (H37Rv, H37Ra), M. avium, M. intracellulare, M. scrofulaceum 균주와 환자의 객담에서 DNA를 추출하여, 383 base pair DNA 양끝의 20 base pair DNA primer (TB-1, -2)와 IS6110 fragment 일부의 DNA 양끝의 20 base pair DNA primer (Sal I-1, -2) 로 PCR을 시행하였으며, 전기 영동후 자외선 발광으로 확인하였다. 결과 : 1) Ethidium bromide 염색후 발광경하에서, Mycobacterium tuberculosis (H37Rv, H37Ra)와 Mycobacterium bovis는 TB-1, -2 primer와 Sal I-1, -2 primer를 이용한 PCR에서 모두 양성을 보였고 Mycobacter intracellulare와 Mycobacterium scrofulaceum은 TB-1, -2 primer를 이용한 PCR에서만 양성을 보였다. 2) Southern Blot 분석에는 두쌍의 primer 모두에서 Mycobacterium tuberculosis (H37Rv, H37Ra)와 Mycobactgerium bovis만이 양성을 나타내었으며 Mycobacterium intracellulare 와 Mycobacterium scrofulaceum은 음성을 나타내었다. 3) Mycobacterium tuberculosis (H37Rv)를 순차적으로 희석하여 시행한 PCR에서 두쌍의 primer 모두에서 Mycobacterium 균 1개체에 해 당되는 1 fg DNA까지 양성을 나타내었다. 4) 임상적으로 진단받은 결핵환자의 시행한, Sal I-1, -2 primer를 이용한 PCR에서 도말 검경 양성군의 객담 29예중 28예인 96.6%에서 양성을 나타내었으며, 도말 정경 음성-배양 양성군에서는 5예중 4예(80.0%), 그리고 도말 검경 음성-배양 음성군에서는 26예중 6예(23.1%)가 양성을 나타내었고 음성 대조군 검체 16예에서 2예(12.5%)에서 양성을 나타내였다. 결론 : 이상의 결과로, PCR은 객담에서의 결핵균의 진단에 있어, 배양과 견줄 수 있는 특이도와 예민도를 보이고 있어, 특정한 경우 진단에 도움을 줄 수 있을 것으로 기대되며, 추후 방법의 개선을 위한 연구가 계속 필요할 것으로 사료된다.

• 대한수의학회지
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• 제41권2호
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• pp.157-165
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• 2001

Amplified fragment length polymorphism(AFLP) technique is based on the polymorphism detection through selective PCR amplification of restriction fragments from digested genomic DNA and thus includes the procedures of the total DNA digestion by endonucleases, ligation of adapters to the ends of the fragments, and following the selective amplification of the restricted DNA fragments. This study were aimed to : (1) determine the genetic variability of S aureus strains, (2) estimate genetic diversity within and among these strains, (3) compare phylogenetic relationships among these strains as genetic markers using AFLP techniques. Genomic DNA was digested with a particular combination of three restriction enzymes with specific recognition sites and the DNA fragments were ligated to restriction specific adapters and amplified using the selective primer combinations. In the S aureus strain, the number of scorable AFLP bands detected per each primer combination varied from 29 to 102, with an average of 61.59 using 27 primer combinations. A total of 1,663 markers were generated, 904 bands of which were polymorphic, showing a 33.48% level of polymorphism with these primer combinations. Among the primer combinations, E02/T02, E02/T03, E04/H02, E02/T01 and E04/H03 primer combinations showed a high level of polymorphism with 0.78, 0.76, 0.74, 0.71 and 0.70, respectively. But T03/H01, E01/T02 and E01/T03 primer combinations showed a low level of polymorphism with 0.38, 0.37 and 0.15, respectively, Therefore, the former primer combinations will be the most effective for AFLP analysis of S aureus. In SA1 sub-types the level of polymorphism of S aureus KCTC 1927 was similar to that of S aureus CU 01(0.825) and higher than those of other strains such as S aureus CU 02 (0.715), S aureus KCTC 2199(0.625), S aureus KCTC 1916(0.607) and S aureus KCTC 1621 (0.553). In SA2 sub-types the level of polymorphism of S aureus CU 07 was similar to that of S aureus CU 08(0.935) and higher than those of both S aureus CU 04(0.883) and S aureus CU 05(0.883) and lower than those of S aureus CU 03(0.583). In SA3 subtypes the level of polymorphism of S aureus CU 11 was similar to that of S aureus CU 12(0.913) and lower than that of S aureus CU 15(0.623). The results proved that AFLP marker analysis of S aureus strain could be used to study the epidemiology of mastitis and in addition, common genotype in geographic region could be useful for the development of an effective vaccine or DNA marker for easy diagnosis of mastitis caused by S aureus infection.

• 식물병연구
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• 제29권1호
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• pp.39-44
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• 2023

To diagnose lily fasciation, lily bulbs showing fasciation were collected from several greenhouses in Jeju Island, South Korea. Bacteria were isolated from the lily bulbs and amplified with both primers for fasA in plasmid and for putative glycosyltransferase epsH gene in chromosome of Rhodococcus fascians. Three bacterial isolates were detected with the P450 primer set and identified as R. fascians by NCBI blast analysis. Twelve bacterial isolates were identified as R. fascians using RS02785 primer set, including the three bacterial isolates identified as the same pathogen using the P450 primer set. Pathogenicity of these bacterial strains identified as R. fascians was demonstrated. Apparent symptoms were observed on wounded lily leaves after inoculation with each bacterial suspension whereas no symptom was found on lily leaves treated with H₂O. Furthermore, bacteria re-isolated from wounded sites were identified as R. fascians. Based on the results, these two sets of primers are recommended for quarantine of R. fascians.

• Restorative Dentistry and Endodontics
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• 제24권1호
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• pp.156-165
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• 1999

The purpose of this study was to evaluate the effect of surface treatments on the shear bond strength between new and old composites. Circular cavities prepared on the center of acrylic resin mold and the prepared cavities were filled with composite resin. They randomly assigned into control group and 8 groups according to the difference in surface treatments of old composites; Control group: no surface treatment, Group 1: surface treated with #120 SiC paper & bonding agent, Group 2: surface treated with #400 SiC paper & bonding agent, Group 3: surface treated with #120 SiC paper, 32% $H_3PO_4$ & bonding agent, Group 4: surface treated with #400 SiC paper, 32% $H_3PO_4$ & bonding agent, Group 5: surface treated with #120 SiC paper, primer & bonding agent, Group 6: surface treated with #400 SiC paper, primer & bonding agent, Group 7: surface treated with #120 SiC paper, 32% $H_3PO_4$, primer & bonding agent, Group 8: surface treated with #400 SiC paper, 32% $H_3PO_4$, primer & bonding agent. New composites were applicated on the old composites of experimental groups. The shear bond strengths for the experimental specimen were measured and the results were analyzed by using one way ANOVA. The observations of surface morphology after SiC paper roughening and debonded surface morphology after shear bond strength test were done by SEM. The results were as follows; 1. Shear bond strengths for specimens roughened with #120 SiC paper matching with the particle size of coarse diamond bur were significantly higher than those for the specimens with #400 SiC paper(P<0.05). By SEM, the surface of the specimens roughened with #120 SiC paper was more irregular than the specimens with #400 SiC paper. 2. Shear bond strengths for specimens treated with 32% $H_3PO_4$ etchant, primer, bonding resin were significantly higher than those for specimens treated with 32% $H_3PO_4$ and bonding resin(P<0.05). 3. Shear bond strengths for the specimens treated with 32% $H_3PO_4$ etchant and bonding resin were significantly higher than those for specimens treated with only bonding resin(P<0.05). There was no remarkable change of surface morphology after 32% $H_3PO_4$ etching. 4. It was possible to observe mixed fracture patterns (the cohesive fracture of old composite and the adhesive fracture between old and new composite) in the specimens roughened with #120 SiC paper, but almost adhesive fracture in the specimens roughened with #400 SiC paper.

• The Journal of Advanced Prosthodontics
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• 제10권3호
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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.

• 농업과학연구
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• 제26권2호
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• pp.33-38
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• 1999

재래산양의 유전자원 보존과 유전적 개량을 위하여 재래산양과 수입산양의 genomic DNA의 유전적 다형을 분석하기 위하여 수행되었으며 재래산양과 수입산양의 유전적 판별 분석은 RAPD 기법을 이용하였으며 공시품종은 재래산양 30두, 재래산양 교잡종 10두, 수입산양 10두를 이용하였다. 재래 산양육과 수입 산양육의 판별을 위한 시료는 재래 산양육 10두와 수입 산양육 10두를 이용하였다. 이들로부터 얻어진 결과를 요약하면 다음과 같다. 1. 재래산양, 수입산양 및 재래산양 교잡종으로부터 추출된 genomic DNA는 전기영동에 의해 약 23kb 크기의 DNA을 얻을 수 있었으며 UV spectrophotometer를 이용하여 흡광도 A 260과 A 280의 비율로 측정한 결과, 그 비율이 1.75~2.10의 범위로 순도는 비교적 양호한 결과를 얻었다. 2. 순수 재래산양의 유전자원의 보존을 위한 재래산양의 유전자 감식여부를 탐색하기 위하여 약 110 여종의 random primer를 이용한 RAPD 기법에 의하여 재래산양, 수입산양 및 교잡종의 다형성을 분석한 결과 random primer OPO-19(5'-CAA ACG TCG G-3')를 이용하였을 때 재래산양에서만 396bp에서 band가 나타났으며 수입산양과 교잡종에서는 band가 나타나지 않았다. 3. 또한, 재래 산양육과 수입 산양육의 유전적인 차이를 구명하기 위하니 RAPD 기법에 의한 genomic DNA의 다형성 분석에 있어서도 random primer OPO-19(5'-CAA ACG TCG G-3')를 사용하였을 때 396bp에서 재래 산양육에서는 band가 나타났지만, 수입 산양육에서는 band가 나타나지 않았다.

• Journal of Korean Dental Science
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• 제2권2호
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• pp.5-11
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• 2009

Objectives : This study investigated the hypothesis that the dentin bond strength of self-etching adhesives (SEAs) may be improved by applying a coat of hydrophobic, neutral adhesive resin in addition to SEA. Method and Materials : The bond strengths of two SEAs - Experimental SEA (EX) and Adper Prompt (AP) - were measured with three bonding protocols. The D/E resin of All-Bond 2 was applied as the hydrophobic, neutral adhesive. Clearfil SE Bond (SE, self-etching primer system) and All-Bond 2 (AB, total etching system) were used as references. The following protocols were used: (1) EX1 (EX 1 coat); (2) EX2 (EX 2 coats); (3) EX+ (EX 1 coat + D/E resin); (4) AP1 (AP 1 coat); (5) AP2 (AP 2 coats); (6) AP+ (AP 1 coat + D/E resin); (7) SE (SE primer + SE bond); (8) SE+ (SE primer + D/E resin); (9) AB (etching + AB primer + D/E resin). Filtek Z250 composite resin was built up and the microtensile bond strength (MTBS) values of the specimens were compared. The fractured surfaces were observed using SEM. Results : When SEA was used as self-etching primer and hydrophobic, neutral adhesive was applied as well, MTBS was significantly higher than that when either one coat or two coats of SEA only were used (p < 0.05). Conclusion : The hydrophobic, neutral adhesive improved the integrity of the bonded interface obtained with SEA.

• Journal of Microbiology and Biotechnology
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• 제9권2호
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• pp.184-190
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• 1999

The use of the polymerase chain reaction (PCR) method was described using two sets of primers based on the ceuN gene (JEJ 1 and JEJ 2) which encodes a protein involved in siderophore transport and 16S rRNA gene (pA and pB) for the sensitive and specific detection of enteropathogen Campylobacter jejuni. Six oligonucleotides were utilized in an amplification experiment and PCR products of predicted sizes were generated from whole cells and boiled cell lysates at the same intensity. Two sets of the primer pairs, JEJ and pAB, were specific enough for all C. jejuni strains tested for the direct use of whole cells without DNA extraction or lysis steps. In the PCR using the pAB primer pair, the detection limit, as determined by the ethidium bromide staining of the amplification products on agarose gels, was at the level of $10^1$ bacteria cells or less in both the pure culture and artificially inoculated milk and chicken enrichment samples, whereas the detection limit with the JEJ primer pair was relatively low, i.e. $10^3$ cells or more in the same PCR samples. The PCR method using either a primer JEJ or pAB was both repeatable and specific for the detection of C. jejuni in food. This method is simply completed within 4 h.

• 한국버섯학회지
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• 제12권3호
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• pp.181-186
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• 2014

본 연구는 느티만가닥버섯 20품종 버섯, 잿빛만가닥 3품종 버섯, 땅지만가닥 1품종 및 돌연변이 느티만가닥버섯 20종류의 유전적 변이를 RAPD 방법에 의해 분석하고자 수행하였다. 이들은 한국, 중국, 일본, 대만 등에서 유래한 것들이다. 느티만가닥버섯에 감마선을 조사하여 돌연변이체를 만들었다. 본 연구에 이용한 primer들은 40종류였고 이 중 31종류의 primer등이 반응을 나타내었다. RAPD 결과들을 분석한 결과 이 들 품종으로는 7개의 cluster로 나뉘어 졌다. 돌연변이체들은 유전적으로 유의적 차이가 있는 subgroup으로 나눌 수 있었다. 이상의 결과들은 본 실험에 사용한 primer들과 포자의 감마선 조사 등을 버섯의 신품종 개발에 유용한 도구가 될 것으로 생각된다.

• 한국어병학회지
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• 제34권1호
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• pp.31-38
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• 2021

본 연구는 1996년부터 2010년 사이에 우리나라 양식넙치와 뱀장어에서 분리하여 Edwardsiella tarda로 동정하였던 18 균주에 대하여 생화학적 특성과 RAPD profile을 비교해봄으로써 이전 양식생물의 질병에 관여되었던 Edwardsiella속 세균의 다양성을 알아보고자 하였다. 생화학적 특성으로 볼 때 이들은 citrate 분해, H₂S 그리고 indole 생산 결과에 차이를 보여 4가지 패턴으로 구분되었으나 모두 E. tarda로 동정되었고 숙주에 따른 분리균의 특성으로 구분되지는 않았다. E. tarda 특이 primer인 EDtT로 PCR을 실시한 결과 18 분리균에서는 모두 약 270 bp의 동일한 band가 검출되었으나 비교 균주로 사용된 E. tarda와 E. ictaluri의 type strain에서는 특이 밴드가 검출되지 않았다. 또한 Ready-To-Go-RAPD kit의 primer 5번과 6번으로 실시한 RAPD PCR 결과, 넙치 분리균, 뱀장어 분리균, 그리고 E. tarda와 E. ictaluri type strain에서 band profile이 뚜렷한 차이를 보여 origin에 따른 특징으로 정리될 수 있었으며 병원체 모니터링을 위한 tool의 하나로 개발될 가능성을 보였다.