假病毒载体在新冠病毒研究中的应用.pptVIP

Development and application of SARS-CoV-2 pseudovirus system 重庆医科大学感染性疾病分子生物学教育部重点实验室 汪凯 2020.10.15 Outline Chimeric Virus and Reverse Genetics of SARS-CoV-2 04 Science  18 Jul 2003: Vol. 301, Issue 5631, 294-296 TIME  8 Oct 2020 RNA病毒域 套式病毒目 冠套病毒亚目 冠状病毒科 正冠状病毒亚科 2002 2012 2019 SARS-CoV-2 COVID-19 8437 cases, 813 deaths 1278 cases, 624 deaths 34 804 348 cases, 1 030 738 deaths /emergencies/diseases/novel-coronavirus-2019/situation-reports/ Chapter 28, Figure 28.2 FIELDS Virology, Sixth Edition Emmie de W. Nat Rev Microbiol (2016) Viruses 2015, 7, 1700-1725 Nature Structural Molecular Biology ,2020 Int. J. Mol. Sci. 2020 FP  six-helix bundle  Zumla, A., Nat Rev Drug Discov. (2016) (Fusion) ① ② Cell line TMPRSS2 Cat B/L TMPRSS2 inhibitor Cat B/L inhibitor HEK 293T — + resistant sensitive Vero — + resistant sensitive Calu-3 + — sensitive resistant Caco-2 + + partially R partially R Nature, 2020 Cell,2020 ① PV is significantly safer to handle. ② PV is easier to handle. Replication deficiency:  single round of infection e.g., easy to introduce point mutation for molecular study; reporter gene Front. Immunol. 2015 Reporter: ① GFP ②luciferase Murine Leukemia Virus Vesicular Stomatitis Virus Pseudotype Virus: the genome of one virus is enclosed in an envelope coded for by another virus. （Cosset FL et al (1992) J Virol 66 , 5671） Cell Host Microbe, 2020 SARS-CoV-2 Spike + VSV △G Pseudovirus /articles/s41596-020-0394-5 Votteler et al., 2008 Envelope of the virus of interest; can be supplied in trans; the gag/pol polyprotein that will make the core of the virion; the reporter gene or gene of interest. King et al., 2016 1. Diagnostic Tool Development: e.g. PCR-based assays (positive control), ELISA-based assays; 2. Studying the mechanism of virus-receptor interaction 3. Screening neutralizing antibodies against SA