Baculovirus Expression System을 이용한 Newcastle Disease Virus 내열성 분리주의 F 단백 유전자 발현

• Published in: Journal of bacteriology and virology Vol. 31; no. 2; pp. 163 - 174
• Main Authors: 장경수, Kyung Soo Chang, 김지영, Ji Young Kim, 김석, Suk Kim, 김태용, Tae Yong Kim, 송희종, Hee Jong Song, 전무형, Moo Hyung Jun
• Format: Journal Article
• Language: Korean
• Published: 대한바이러스학회 2001; 대한미생물학회
• Subjects: Expression of F protein gene; Newcastle disease virus; Recombinant baculovirus system; Sf-9 cell; Thermostable Korean isolate; baculovirus system; Newcastle disease virus; Expression of F protein gene; Sf-9 cell; Thermostable Korean isolate; Recombinant
• ISSN: 1598-2467; 2093-0429

The gene (1,707 bp) encoding fusion (F) protein of CBP-1 strain, a heat-stable Newcastle disease virus (NDV) isolated from the diseased pheasant in Korea was amplified by reverse transcription-polymerase chain reaction (RT-PCR) and the clone of pVL-NDF inserted with NDV F gene was constructed. Using Baculo Gold Transfection kit, pVL-NDF plasmid and linear baculovirus DNA were cotransfected into Sf-9 cells, and F gene in NDVF recombinant baculovirus was identified by restriction enzyme cleavage patterns and Southern blot hybridization. The Sf-9 cells infected with the NDVF recombinant baculovirus showed the typical cytopathic effects with syncytial cell formation. The properties of the expressed F protein were examined by indirect fluorescent assay, immunocytochemistry and indirect dot-immunoassay. By SDS-PAGE, the expressed F protein with the size of 55 kDa was detected in the cells infected for 5 days with 0.125 to 4.0 moi of the recombinant baculovirus and in the cells infected with 2 moi of the recombinant baculovirus for 1 to 6 days. The guinea pigs inoculated with the mixtures of the infected Sf-9 cell cultures and adjuvants showed significant immunoreactivity, as measured by indirect enzyme-linked immunosorbent assay.