A transgenic Marc-145 cell line of piggyBac transposon-derived targeting shRNA interference against porcine reproductive and respiratory syndrome virus.
Porcine reproductive and respiratory syndrome (PRRS) is now considered to be one of the most important diseases in countries with intensive swine industries. The two major membrane-associated proteins of porcine reproductive and respiratory syndrome virus (PRRSV), GP5 and M (encoded by ORF5 and ORF6 genes, respectively), are associated as disulfide-linked heterodimers (GP5/M) in the virus particle. In this study, we designed 5 of the small hairpin RNAs (shRNAs) targeting the GP5 and M gene of PRRSV respectively, and investigated their inhibition to the production of PRRSV. The highest activity displayed in shRNAs of the ORF6e sequence (nts 261-279), which the inhibition rate reached was 99.09%. The result suggests that RNAi technology might serve as a potential molecular strategy for PRRSV therapy. Furthermore, the transgenic Marc-145 cell line of piggyBac transposon-derived targeting shRNA interference against PRRS virus was established. It presented stable inhibition to the replication and amplification of PRRS. The work implied that shRNAs targeting the GP5 and M gene of PRRSV may be used as potential RNA vaccines in vivo, and supplied the screening methods of transformed pig embryonic fibroblast which are prerequisite for the disease-resistant transgenic pigs to PRRS.
Key Laboratory of Animal Virology of Ministry of Agriculture, College of Animal Sciences, Zhejiang University, Hangzhou 310058, People’s Republic of China.
Fang
F
Zhou
Shuang
S
Liang
An-hui
AH
Chen
Chabungbam Orville
CO
Singh
Roy
R
Bhaskar
Yan-shan
YS
Niu
Yun-gen
YG
Miao
D000818:Animals; D002460:Cell Line; D003001:Cloning, Molecular; D015967:Gene Expression Regulation, Viral / Q000502:physiology; D010957:Plasmids; D019316:Porcine respiratory and reproductive syndrome virus / Q000502:physiology*; D034622:RNA Interference / Q000502:physiology*; D034741:RNA, Small Interfering / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013552:Swine; D014162:Transfection; D014759:Viral Envelope Proteins / Q000235:genetics / Q000378:metabolism*
D034741:RNA, Small Interfering; D014759:Viral Envelope Proteins; C114789:glycoprotein 5, PRRSV