Baculovirus를 利用한 Canine Parvovirus VP2蛋自質의 發現 = Expression of Canine Parvovirus VP2 by a Recombinant Baculovirus

국문 초록 (Abstract)

국내에서 分離된 개파보바이러스주(V20주)의 VP2 遺傳子를 baculovirus system을 이용하여 발현시켜 다음의 結果를 얻었다.
1. 개파보바이러스의 VP2 유전자를 PCR에 의해 增幅하여 1755bp의 VP2遺傳子를 pUC19에 클로닝하여, 클로닝된 遺傳子를 polyhedrin promoter를 가지는 baculovirus expression vector인 pVL1393에 옮겨 VP2 발현벡터인 pVL1393-VP2를 얻을 수 있었다.
2. pVL1393-VP2 plasmid와 baculovirus DNA와의 homologous recombination에 의해 재조합바이러스인 VP2-BV를 얻을 수 있었으며, 그 발현효율은 2.000-5.000 HAU/0.05 ml이었다.
3. 免疫沈澱法에 의해 발현된 단백질은 개파보바이러스의 VP2단백질과 유사한 64 kb에 달하는 것이었으며, 血球凝集能을 지니고 있었다
4. 血球凝集能을 지닌 蛋白質이 여러 陽性血淸에 있어서 抗體수준을 測定할 수 있는지 개파보바이러스항원과의 相關性을 比較한 바 0.94 (n=125. p<0.01)의 相關係數를 보였다
5. 발현 VP2白은 virus-like particles를 形成하였으며, 그 크기로는 개파보바이러스와 비슷한 25 ㎚의 크기를 갖았다.

다국어 초록 (Multilingual Abstract)

Canine parvovirus(CPV) is a member of autonomous replicating parvoviruses and is aetiologically associated with enteritis and mycoarditis in puppies. The capsids of CPV are composed of three structural proteins: VP1, VP2 and VP3. The VP2 protein is the major component of capsid. The VP2 gene of a canine parvovirus. V20 strain isolated in Korea was cloned into baculovirus expression vector, and subsequently the VP2 protein was expressed by a recombinant baculovirus under the control of polyhedrin promoter. The recombinant VP2 protein expressed in Spodoptera frugiperda 9(Sf9) cells was detected by haemagglutination(HA) test and immunofluorescent antibody assay. Molecular weight of the recombinant VP2 protein expressed was estimated as 64Kd when tested by immunoprecipitation test using anti-CPV monoclonal antibody. In haemagglutination inhibition(Ⅲ) test. 8 HA units of the recombinant VP2 protein antigen was successfully utilized to determine a level of antibody against CPV in various positive sera. The recombinant VP2 protein showed also the capability to form virus like particles similar in size and appearance to the CPV virions.