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In this study, a DNA-launched reverse genetics system was developed from a type 2 porcine reproductive and respiratory syndrome virus (PRRSV) strain, KNU-12. The complete genome of 15,412 nucleotides was assembled as a single cDNA clone and placed under the eukaryotic CMV promoter. Upon transfection of BHK-tailless pCD163 cells with a full-length cDNA clone, viable and infectious type 2 progeny PRRSV were rescued. The reconstituted virus was found to maintain growth properties similar to those of the parental virus in porcine alveolar macrophage (PAM) cells. With the availability of this type 2 PRRSV infectious clone, we first explored the biological relevance of ORF5a in the PRRSV replication cycle. Therefore, we used a PRRSV reverse genetics system to generate an ORF5a knockout mutant clone by changing the ORF5a translation start codon and introducing a stop codon at the 7th codon of ORF5a. The ORF5a knockout mutant was found to exhibit a lack of infectivity in both BHK-tailless pCD163 and PAM-pCD163 cells, suggesting that inactivation of ORF5a expression is lethal for infectious virus production. In order to restore the ORF5a gene-deleted PRRSV, complementing cell lines were established to stably express the ORF5a protein of PRRSV. ORF5a-expressing cells were capable of supporting the production of the replicationdefective virus, indicating complementation of the impaired ORF5a gene function of PRRSV in trans.
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참고문헌 (23)
참고문헌 신청
[학술지(정기
Albina E / 1997 / Epidemiology of porcine reproductive and respiratory syndrome (PRRS): an overview / Vet. Microbiol 55 : 309 ~ 316
Bautista EM / 2002 / Functional properties of the predicted helicase of porcine reproductive and respiratory syndrome virus / Virology 298 : 258 ~ 270
Benfield DA / 1992 / Characterization of swine infertility and respiratory syndrome (SIRS) virus (isolate ATCC VR-2332) / J. Vet. Diagn. Invest 4 : 127 ~ 133
Cavanagh D / 1997 / Nidovirales: a new order comprising coronaviridae and arteriviridae / Arch. Virol 142 : 629 ~ 633
Albina E / 1997 / Epidemiology of porcine reproductive and respiratory syndrome (PRRS): an overview / Vet. Microbiol 55 : 309 ~ 316
Bautista EM / 2002 / Functional properties of the predicted helicase of porcine reproductive and respiratory syndrome virus / Virology 298 : 258 ~ 270
Benfield DA / 1992 / Characterization of swine infertility and respiratory syndrome (SIRS) virus (isolate ATCC VR-2332) / J. Vet. Diagn. Invest 4 : 127 ~ 133
Cavanagh D / 1997 / Nidovirales: a new order comprising coronaviridae and arteriviridae / Arch. Virol 142 : 629 ~ 633
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