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- 저자 : Taniguchi Koki) (검색결과 4 건)
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JBR(Jeju Island Bovine Rotavirus)의 Sequencing에 의한 Homology 비교에 관한 연구
이종익,谷口孝喜,김응률,류영수,박선오,송진욱,유제현,조홍찬,차광종,浦澤正三,이태협,김유성,이영건 건국대학교 동물자원연구센터 1998 動物資源硏究誌 Vol.19 No.-
본 연구는 PCR에 의해 로타바이러스의 serotype을 결정하고 염기배열의 유사성을 다른 지역에서 발견된 로타바이러스와 비교 분석하였으며 그 결과는 다음과 같다. 1. 제주도 목장에서 송아지 설사변을 채취하여 로타바이러스를 분리 후 MA104세포에 감염시켜 세포변성 효과(CPE)를 확인하였다. 2. 전자현미경에 의해 형태학적로 로타바이러스임이 판명되었으며 형광항체법에 의해 MA104세포에 감염된 것을 재확인 하였다. 3. 제주도 송아지 로타바이러스(JBR)RNA를 PAGE에 의해 genotype를 분석한 결과 bovine특유의 4:2:3:2 pattern이지만 일반 PAGE로 분석한 결과는 NCDV, UK, KK3, A5-37, 61A, B223와 차이가 있었으며, Ns-5, Nc-5, Kawatabi(Japan)와는 비슷한 경향을 보였다. 4. Plaquing 후 titer한 결과 NCDV, UK보다는 낮은 ??PFU/ml이었다. 5. RNA-RAN hybridization과 ELISA 및 VP7과 VP4의 1차, 2차 PCR 사물을 1% agarose(EtBr 1㎕의 TAE)에서 전기영동한 결과 G6P11의 serotype이었다. 6. JBR의 P serotype이 동일하게 나타난 B223과의 비교 결과 총 JVP8 750 bases 중에서 731개의 염기가 B223과 같은 것으로 나타나 염기배열의 유사성은 97.47%로 나타났고, 아미노산 배열의 유사성은 97.57%로 나타났다. This study was carried our to identify JBR's serotype by polymerase chain reaction(PCR) and to analyze homology of JBR' sequence. The results obtained in these experiments were as follows: 1. Fecal samples of calf diarrhea were taken on farms in Jeju island, rotavirus was isolated and cytopathic effect(CPE) was determined after infection to MA104 cell. 2. Morphological evaluation on electron microscopy proved it as rotavirus. Also, its infection to MA104 cell was reidentified using a fluorescence antibody method. 3. Genotype of Jeju island bovine rotavirus(JBR) analyzed thorough PAGE was 4:2:3:2 pattern, which was unique in bovine and that analyzed through general PAGE was somewhat different from NCDV, UK, KK3, A5-37, 61A, B223 and similar to Nstool-5, N cultrue-5 and Kawatabi(Japan). 4. By titration after plaquing, the level was ??PEU/ml, which was lower than those of NCDV and UK. 5. Electrophoresis analysis of RNA-RNA hybridization. ELISA, adn first and second PCR products of VP7 and VP4 in 1% agarose(TAE+1㎕ EtBr) revealed that the rotavirus was a serotype of G6P11. 6. Alignment of two(JBR and B223) amino acid sequences is 97.57% and nucleotide sequences is 97.47%.
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치즈 제조 부산물을 이용한 ELISA Blocking Agent의 개발에 관한 연구
유제현,송진욱,조흥찬,차광종,박범석,김응률,이종익,이중복,谷口孝喜 건국대학교 동물자원연구센터 1999 動物資源硏究誌 Vol.20 No.-
본 연구는 유청과 탈지분유를 blotto로 이용하여 ELISA 수행시 항원·항체반응에 비특이반응을 막는 Blocking agent를 탐색하기 위해 실험을 수행했다. MAb의 역가측정 결과 유의차가 인정되는 OD값이 0.3이상 검색된 희석배율은 1B2의 경우 819,200배, 6D4의 경우 1,638,400배, 15B10의 경우 204,800배이었다. 서로 다른 Blotto가 항원·항체반응에 미치는 영향을 각기다른 MAb를 이용하여 조사해 본 결과 MAb Yo-156, Yo-5는 Blocking agent로 표준적으로 사용하고 있는 BAS가 비교하여 group A 바이러스 7종 모두에 비슷한 정도의 발색을 일으켰으며, MAb G6는 G6 type바이러스 4종 모두에서 비슷한 정도의 발색을 일으켰고, MAb G10은 G10 type 바이러스 3종 모두에서 비슷한 정도의 발색을 일으켰다. 따라서 본 실험에 사용한 모든 종류의 blotto소재인, 탈지분유1(서울우유(협)), 탈지분유2((주)매일유업), 혼합탈지분유((주)매일유업), whey powder1(Sigma), whey powder2 ((주)삼익유가공), WPC34((주)환이상사), WPC800((주)후드피아), WPC8000 ((주)후드피아), WPI((주)환이상사)는 현재 Blocking agent로 사용하고 있는 BSA와 비교하여 유사하게 비특이 반응을 막아주는 것으로 밝혀져 경제적으로 보다 저렴한 blotto 소재로 이용이 가능하다는 사실을 확인하였다. Enzyme-Linked-Immuno-Sorbent-Assay (ELISA) has been frequently used for detection of interaction of antigen and antibody. Many efforts have been made to reduce nonspecific reaction of ELISA. Bovine serum albumin (BSA) is usually used for blocking the nonspecific reaction even thought it is expensive. Recently skin milk has been focused on blocking nonspecific reaction of ELISA. This study aimed to substitute expensive blocking agent of ELISA with cheap by-product of cheese manufacture. To evaluate the substitution of BSA with wheys made in different companies, ELISA was carried out using monoclonal antibodies (MAbs) against rotavirus. The blocking capacity of the whey was compared with that of the BSA. The commercial wheys showed the same activity as BSA when ELISA was performed using MAb YO-156, S2-37, and Yo-5. To further investigate whether the whey affects the detection specificity of MAb, the ELISA was carried out using type specific MAbs. MAb G6 specific to serotype G6 bound to only rotavirus serotype G6 such as NCDV, UK, JBR, and Jedong strain whereas MAb G10 specific to serotype G10 bound to serotype G10 rotavirus only, demonstrating that the whey a and no effect on the binding specificity of MAb with respect to antigen-antibody reaction. These data indicate that the whey may have no effect on specific interation of antigen-antibody and may be widely used for blocking substitute because of competitive priority of its price.
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차광종,송진욱,조홍찬,김용휘,유대열,이중복,이종기,곡구효보,유제현,Cha, Kwang-Jong,Song, Jin-Ook,Cho, Hong-Chan,Kim, Yong-Hee,Yu, Dae-Yeul,Lee, Joong-Bok,Lee, Chong-Kee,Koki, Taniguchi,Yu, Jae-Hyeun 대한미생물학회 1999 Journal of Bacteriology and Virology Vol.28 No.1
Rotaviruses belong to Reoviridae causes diarrhea in human beings as well as domestic animals. This study was conducted to see what type of human rotaviruses are distributed in Seoul and Kyung-gi province. Twenty two of 81 patients showed rotavirus positive with diagnostic kit and RNA electropherosis. We isolated all of rotaviruses from the patients. Electropherotypes of 22 isolates showed 4:2:3:2 pattern whereas those migration patterns were long type. All of those isolates belonged to group A. Twenty out of 22 isolates reacted with monoclonal antibodies specific to G1, P1A and subgroup II, whereas rest of them, A-29 and K-30 reacted with subgroup I specific monoclonal antibody. The nucleotide sequence of an isolate K-21 showed $98{\sim}100%$ and $90{\sim}96%$ homologies with those of Wa and KU strain, respectively.
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차광종(Gwang Jong Cha),송진욱(Jin Wook Song),조홍찬(Hong Chan Jo),김용휘(Yong Hwi Kim),유대열(Dae Yeol Yoo),이중복(Joong Bok Lee),이종기(Jong Gee Lee),유제현(Je Hyun Yoo),Taniguchi Koki 대한바이러스학회 1999 Journal of Bacteriology and Virology Vol.29 No.2
Rotaviruses belong to Reoviridae causes diarrhea in human beings as well as domestic animals. This study was conducted to see what type of human rotaviruses are distributed in Seoul and Kyung-gi province. Twenty two of 81 patients showed rotavirus positive with diagnostic kit and RNA electropherosis. We isolated all of rotaviruses from the patients. Electropherotypes of 22 isolates showed 4:2:3:2 pattern whereas those migration patterns were long type. All of those isolates belonged to group A. Twenty out of 22 isolates reacted with monoclonal antibodies specific to G1, P1A and subgroup II, whereas rest of them, A-29 and K-30 reacted with subgroup I specific monoclonal antibody. The nucleotide sequence of an isolate K-21 showed 98-100% and 90-96% homologies with those of Wa and KU strain, respectively.
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