[Construction of an infectious clone of equine infectious anemia virus by N-glycosylation reverse-mutations].
To elucidate the role of N-glycosylation in fetal donkey dermal cell (FDD)-attenuated equine infectious anemia virus (EIAV), we constructed an N-glycosylation reverse-mutation molecular clone, pLGN191N236N246. This viral molecular clone was derived from the infectious clone pLGFD3-8 by site-directed mutagenesis. This clone was used to transfect fetal donkey dermal (FDD) cells. Infectious characteristics of transfectants were monitored by RT-PCR, indirect immune fluorescence and reverse transcriptase activity assay. After three passages in FDD cells, viral replications in the supernatant of cell cultures were detected by all the above three methods and viral particles were clearly observed by electron microscopy. The construction of the N-glycosylation reverse-mutation infectious clone provides a solid basis for further study of the mechanism of attenuated pathogenesis and increased immune protection of EIAV attenuated vaccines.
Han
Xiu'e
X
National Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China. hanxiue_2005@163.com
Quan
Yanping
Y
Gao
Xu
X
Xiang
Wenhua
W
Zhou
Jianhua
J
D000818:Animals; D002460:Cell Line; D019084:Fluorescent Antibody Technique, Indirect; D005818:Genetic Engineering / Q000379:methods*; D006023:Glycoproteins / Q000235:genetics; D006031:Glycosylation; D004860:Infectious Anemia Virus, Equine / Q000235:genetics* / Q000302:isolation & purification / Q000502:physiology*; D008854:Microscopy, Electron; D016296:Mutagenesis; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012867:Skin / Q000378:metabolism / Q000648:ultrastructure / Q000821:virology; D014162:Transfection; D014759:Viral Envelope Proteins / Q000235:genetics; D014779:Virus Replication
D006023:Glycoproteins; C091940:SU protein, equine infectious anemia virus; D014759:Viral Envelope Proteins; D012194:RNA-Directed DNA Polymerase