Construction of JSRV-NM Pseudovirions by High Efficiency Packaging System of the Lentivirus
10.3969/j.issn.0253-9896.2014.08.004
- VernacularTitle:基于慢病毒载体系统构建重组JSRV-NM假病毒
- Author:
Shumin GONG
;
Guangming LI
;
Zhimin WU
;
Lizhen DONG
;
Bin CHENG
;
Bin ZHANG
;
Ze ZHU
- Publication Type:Journal Article
- Keywords:
JSRV-NM viruses;
pseudovirions;
lentivirus vector;
env gene;
virus packageing
- From:
Tianjin Medical Journal
2014;(8):749-751
- CountryChina
- Language:Chinese
-
Abstract:
Objective To overcome the fact that SRV-NM virus can only multiple and amplify through partially pu-rified jaagsiekte retrovirus inoculated intratracheally in sheep but it cannot be augmented using in vitro cell culture, we con-structed JSRV-NM pseudovirions based on high efficiency packing system of lentivirus. Methods Lentivirus of three high efficiency packing plasmids system pMD.G, pCMV-HIV 8.2 and pHIV-eGFP was developed, and JSRV-NM-env coated plasmid pCMVJSRV-NM was used to substitute VSV-G virus coated plasmid pMD.G then co-transfected into 293T cells to replicate, package and produce restructured JSRV-NM pseudovirions. Gene expression of pseudovirion was determined through WPRE using real time PCR; Virus infectivity was detected through inoculating JSRV-NM pseudovirions into 24 pore plates. Results We construct JSRV-NM pseudovirions successfully based on the lentivirus system. JSRV-NM pseudo-virions can also be concentrated to higher titer (108 TU/mL detected by real time PCR by ultracentrifugation without signifi-cant loss of activity. JSRV-NM and VSV-G pseudovirions infected on Hela cells (both MOI= 3) respectively and no obvi-ous difference were shown on their infection efficiency detected by real time PCR. Conclusion Based on lentivirus system, JSRV-NM pseudovirions can be multipled and amplified in 293T cell culture in vitro. JSRV-NM pseudovirions is stable without loss its infection activity and the requirements of biological laboratory safety II was also met. JSRV-NM pseudoviri-ons will provide a useful tool for further study of JSRV-NM-env infection across species or its induction of lung adenocarci-noma.
