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국내분리 돼지 뇌심근염바이러스의 poly(C)-tract의 다형태성
현방훈,김효진,김인중,표현미,김선미,김성희,김재조,임성인,송재영,Hyun, Bang-Hun,Kim, Hyo-Jin,Kim, In-Joong,Pyo, Hyun-Mi,Kim, Sun-Mi,Kim, Seong-Hee,Kim, Jae-Jo,Lim, Seong-In,Song, Jae-Young 대한수의학회 2010 大韓獸醫學會誌 Vol.50 No.3
Encephalomyocarditis virus (EMCV) belongs to the genus Cardiovirus within the family Picornaviridae. EMCV has been recognized either as a cause of mortality in young pigs, due to acute myocarditis, or of reproductive failure in sows. An EMCV K3 strain was isolated from the heart and brain in a mummified and aborted swine fetus in 1989. For the molecular characterization of the poly(C)-tract of EMCV Korean isolates, K3 strain, viral RNA was extracted and digested with RNase T1, and analyzed the length of the poly(C)-tract by polyacrylamide gel electrophoresis. The poly(C) regions also were amplified by RT-PCR and sequenced. The present study shows that K3 strain of EMCV had a short polymorphic poly(C) tracts (5 to 30 C's) with sequences consisting of $C_9$, $C_{10}$, $C_{13}$, $C_{14}$, $C_{16}$, $C_{20}$, $CUC_{11}$, $C_8UCUC_3UC_{10}$, $C_9UCUC_3UC_{10}$, $C_{10}UCUC_3UC_{10}$, etc. These polymorphism of poly(C)-tracts of EMCV K3 strain implies the historical information of in vivo and/or in vitro passage.
현방훈,김인중,표현미,차상호,박지연,송재영,조인수,양창범,안수환,이중복,Hyun, Bang-Hun,Kim, In-Joong,Pyo, Hyun-Mi,Cha, Sang-Ho,Park, Ji-Yeun,Song, Jae-Young,Cho, In-Soo,Yang, Chang-Bum,An, Soo-Hwan,Lee, Joong-Bok 대한수의학회 2009 大韓獸醫學會誌 Vol.49 No.1
The molecular genetic characterization of Aujeszky's disease virus (ADV) Yangsan strain (ADVYS), a Korean isolate, was investigated by analyzing the electrophoresis patterns and the physical maps of the viral DNA digested with various endonucleases. To establish DNA library for ADV-YS, twelve major BamHI restricted segments were cloned. Each location of the segments in the ADV genome was determined by sequence comparison with the sequences reported in Genbank and those sequences of the both termini of the segments. Physical maps were constructed based on the electrophoresis patterns of the digested viral DNA by restriction endonuclease and the results of Southern blot analyses with various DIG labeled probes originated from those of enzyme restricted segments of virulent (Shope) and avirulent (Bartha) strain. Comparing ADV-YS with a standard strain of Kaplan in the maps of restriction enzymes, following major respects were identified: (i) disappearance of BamHI restriction site between the first and second BamHI segments, (ii) creation of the BamHI restriction site in the fifth segment, and (iii) generation of the BglII site in the unique short (US) region. The genome of ADV-YS also contains a type 2 herpesvirus DNA molecule (in which the US region only inverts itself relative to the unique longregion) like all other ADV strains except Norden strain(type3), analyzed up to date. The size of the ADV genome estimated from the sizes of the restriction enzyme fragments, was approximately 145.3 kb (BamHI) or 145.4 kb (BglII). BamHI enzyme cleavage patterns were compared among the five Korean ADV isolates: Yangsan, Yongin, Dangjin, Jincheon and Iksan strains. Difference either in the number or in the size of the DNA fragments, suspected regions of termini of IR and TR, could be detected among all five strains.
Current situation, diagnosis, prevention & control of PED and TGE (PED와 TGE의 병태, 진단 및 방역전략)
현방훈 대한수의학회 2007 대한수의학회 학술대회발표집 Vol.2007 No.-
Porcine epidemic diarrhea (PED) and transmissible gastroenteritis (TGE) are very similar in pathogenetic features and are highly contagious, enteric viral diseases of swine characterized by severe diarrhea and a high mortality in piglets. In Korea, TGE outbreaks have decreased coincident with the spread of porcine respiratory coronavirus (PRCV), but PED outbreaks however have been reported continuously. PED is a major devastating enteric disease and one of the important diseases should be controlled intensively with PRRS and PMWS in Korean pig industry. The reviewed here are the current situation, Diagnosis, prevention and control for PED and TGE in Korea. As the control measures to PED, recently studies on immunoprophylaxis using PEDV neutralizing scFv and vaccine enhanced by viral surface-expressed Fc molecules are introduced. Several requirements to PED control have been proposed to account for current PED status in Korea. First, enhancement of efficacy or immunogenicity of the current vaccines. Second, active guide on vaccine program and biosecurity to pig breeders, especially the farm in enzootic status. Third, Analysis of the economic assessment for PED outbreaks. Fourth, encouragement of basic research on pathogenesis and mechanism of the interaction between the virus and the host cell. Fifth, development of new measures for the disease control of persistent infection in a farm. Sixth, standardization of the diagnostic assay, especially serologic test. Seventh, establishment of cell lines to stably support wild-type PEDV replication and serial viral propagation, and evaluation of the neutralizing activity of vaccine-induced antibodies to PEDV field strains in swine.
Baculovirus 를 이용한 Aujeszky's Disease Virus gIII 단백질 발현
송재영,이중복,현방훈,박종현,김병한,권창희,전무형,안수환 대한바이러스학회 1992 Journal of Bacteriology and Virology Vol.22 No.2
The gIII gene located in U region of Yangsan strain, a field isolate of Aujeszkys disease virus (ADV) in Korea, was cloned into pTZ18R and sequenced. The glll gene consisting of 1,437 nucleotides showed 98 % sequence homology with that of Becker strain, a reference strain of ADV. The gene encoding gK of Yangsan strain was placed under the control of Autographa californica nucleopolyhedrosis virus (AcNPV) polyhedrin promoter, and expressed by the derived recombinant baculovirus using Spodoptera frugiperda 9 (Sf9) cells. The expressed g Ill was a protein with molecular weight of 72kd determined by immunoprecipitation and Western blotting assay using anti-ADV polyclonal antibodies and anti-g IE monoclonal antibody. The partially purified g lll protein was utilized as antigen in the radial immunodiffusion enzyme assay (RIDEA) to detect the specific antibody against ADV in pig sera. The results indicated that the sensitivity of RIDEA with the recombinant gIll protein antigen (98% ) was as high as that with the conventional glycoprotein antigen extracted from the ADV infected cells. In addition, the false positive and false negative reactions in gIE RIDEA were significantly reduced than the conventional glycoprotein RIDEA as judged from the results of standard serum neutralization test.
Baculovirus를 이용한 Aujeszky's Disease Virus gⅢ 단백질 발현
송재영,이중복,현방훈,박종현,김병한,권창희,전무형,안수환 충남대학교 생물공학연구소 1993 생물공학연구지 Vol.3 No.-
The g Ⅲ gene located in U_L region of Yangsan strain, a field isolate of Aujeszky's disease virus (ADV) in Korea, was cloned into pTZ18R and sequenced. The gⅢ gene consisting of 1,437 nucleotides showed 98% sequence homology with that of Becker strain, a reference strain of ADV. The gene encoding gⅢ of Yangsan strain was placed under the control of Autographa californica nucleopolyhedrosis virus (AcNPV) polyhedrin promoter, and expressed by the derived recombinant baculovirus using Spodoptera frugiperda 9 (Sf9) cells. The expressed gⅢ was a protein with molecular weight of 72kd determined by immunoprecipitation and Western blotting assay using anti-ADV polyclonal antibodies and anti-gⅢ monoclonal antibody. The partially purified gⅢ protein was utilized as antigen in the radial immunodiffusion enzyme assay (RIDEA) to detect to specific antibody against ADV in pig sera. The results indicated that the sensitivity of RIDEA with the recombinant gⅢ protein antigen (98%) was as high as that with the conventional glycoprotein antigen extracted from the ADV infected cells. In addition, the false positive and false nagative reactions in gⅢ RIDEA were significantly reduced than the conventional glycoprotein RIDEA as judged from the results of standard serum neutralization test.