Affiliations
- College of Life Sciences, Henan Normal University, Xinxiang 453007, China; Beijing Institute of Radiation Medicine, Beijing 100850, China.
- Beijing Institute of Radiation Medicine, Beijing 100850, China.
- Beijing Institute of Radiation Medicine, Beijing 100850, China; Cooperative Innovation Center for Molecular Target New Drug Study, University of South China, Hengyang 421001, China.
- Beijing Institute of Radiation Medicine, Beijing 100850, China. Electronic address: xjwang@bmi.ac.cn.
- College of Life Sciences, Henan Normal University, Xinxiang 453007, China; Beijing Institute of Radiation Medicine, Beijing 100850, China. Electronic address: sqwang@bmi.ac.cn.
PMID: 34182038 PMCID: PMC8233049 DOI: 10.1016/j.jviromet.2021.114221
Abstract
SARS-CoV-2 is the culprit causing Coronavirus Disease 2019 (COVID-19). For the study of SARS-CoV-2 infection in a BSL-2 laboratory, a SARS-CoV-2 pseudovirus particle (SARS2pp) production and infection system was constructed by using a lentiviral vector bearing dual-reporter genes eGFP and firefly luciferase (Luc2) for easy observation and analysis.
Comparison of SARS2pp different production conditions revealed that the pseudovirus titer could be greatly improved by: 1) removing the last 19 amino acids of the spike protein and replacing the signal peptide with the mouse Igk signal sequence; 2) expressing the spike protein using CMV promoter other than CAG; 3) screening better optimized spike protein sequences for SARS2pp production; and 4) adding 1 % BSA in the SARS2pp production medium.
For infection, this SARS2pp system showed a good linear relationship between MOI 2-0.0002 and then was successfully used to evaluate SARS-CoV-2 infection inhibitors including recombinant human ACE2 proteins and SARS-CoV-2 neutralizing antibodies. The kidney, liver and small intestine-derived cell lines were also found to show different susceptibility to SARSpp and SARS2pp.
Given its robustness and good performance, it is believed that this pseudovirus particle production and infection system will greatly promote future research for SARS-CoV-2 entry mechanisms and inhibitors and can be easily applied to study new emerging SARS-CoV-2 variants.
Keywords: Pseudovirus; SARS-CoV-2; Susceptible cell lines; Viral entry inhibitors.
