小反刍兽疫病毒感染性cDNA克隆的构建与病毒拯救

doi:10.11843/j.issn.0366-6964.2023.07.026

Abstract

Abstract: Peste des petits ruminants virus (PPRV) is an important pathogen affecting the global livestock industry. The purpose of this research is to establish a stable and reliable PPRV reverse genetic manipulation platform, to provide an effective technical platform for the analysis of PPRV pathogenic mechanism, immune escape mechanism, virus replication mechanism and other basic theoretical research, at the same time, it will lay the necessary preliminary foundation for the development of safer and more effective new vaccines. The RNA of the PPRV Clone9 strain was extracted, and the PPRV antigenome cDNA was amplified by RT-PCR in six sections. The full-length antigenome of the PPRV Clone9 strain was digested and ligated to obtain pB-PPRV, and the PPRV nucleoprotein (N), three helper plasmids for phosphoprotein (P) and large protein (L). pB-PPRV and helper plasmids were co-transfected into BHK-T7 cells mediated by liposomes. Three days after transfection, Vero cells were infected by freezing and thawing three times. Significant cytopathic changes (CPE) were observed in the second generation. The results of indirect immunofluorescence, Western blot, RT-PCR and sequence determination showed that the infectious virus was rescued. The rescue virus (rPPRV-Clone9) was stably passaged on Vero cells, and the proliferation dynamics were similar to the parental virus. In this study, the reverse genetics system of PPRV Clone9 strain was successfully established.

Key words: peste des petits ruminants virus, reverse inheritance, virus rescue

CLC Number:

S852.65

WANG Yu, GAO Yueyi, GAO Jinyuan, LIU Weijie, XU Huilin, XUE Qinghong. Establishment of Reverse Genetics System of Peste des petits Ruminants Virus Clone9 Strain[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(7): 2956-2963.

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Establishment of Reverse Genetics System of Peste des petits Ruminants Virus Clone9 Strain

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