Welcome To Website IAS

Hot news
Achievement

Independence Award

- First Rank - Second Rank - Third Rank

Labour Award

- First Rank - Second Rank -Third Rank

National Award

 - Study on food stuff for animal(2005)

 - Study on rice breeding for export and domestic consumption(2005)

VIFOTEC Award

- Hybrid Maize by Single Cross V2002 (2003)

- Tomato Grafting to Manage Ralstonia Disease(2005)

- Cassava variety KM140(2010)

Centres
Website links
Vietnamese calendar
Library
Visitors summary
 Curently online :  68
 Total visitors :  7666693

Analysis of SARS-CoV-2 infection dynamic in vivo using reporter-expressing viruses
Saturday, 2021/10/16 | 07:17:10

Chengjin Ye, Kevin Chiem, Jun-Gyu Park, Jesus A. Silvas, Desarey Morales Vasquez, Julien Sourimant, Michelle J. Lin, Alexander L. Greninger, Richard K. Plemper, Jordi B. Torrelles, James J. Kobie, Mark R. Walter, Juan Carlos de la Torre, and Luis Martinez-Sobrido

 

PNAS October 12, 2021 118 (41) e2111593118

Significance

To date, due to the insufficient expression level of reporter genes from previous recombinant (r)SARS-CoV-2, in which the viral open reading frame (ORF) 7a gene was replaced by reporter genes, tracking the SARS-CoV-2 infection dynamic has been challenging. Here, we engineered rSARS-CoV-2 expressing fluorescent (Venus) or luciferase (Nano luciferase) reporter genes from the viral nucleocapsid (N) locus, without deleting any viral gene. These novel reporter-expressing rSARS-CoV-2, which give a higher expression level of reporter genes, allow us to monitor SARS-CoV-2 replication dynamic both in vitro and in vivo. These new reporter-expressing rSARS-CoVs-2 represent an excellent option to assess viral replication, tropisms, and pathogenicity as well as the rapid in vivo evaluation of effective countermeasures for the treatment of SARS-CoV-2 infection.

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of the current COVID-19 pandemic, is one of the biggest threats to public health. However, the dynamic of SARS-CoV-2 infection remains poorly understood. Replication-competent recombinant viruses expressing reporter genes provide valuable tools to investigate viral infection. Low levels of reporter gene expressed from previous reporter-expressing recombinant (r)SARS-CoV-2 in the locus of the open reading frame (ORF)7a protein have jeopardized their use to monitor the dynamic of SARS-CoV-2 infection in vitro or in vivo. Here, we report an alternative strategy where reporter genes were placed upstream of the highly expressed viral nucleocapsid (N) gene followed by a porcine tescherovirus (PTV-1) 2A proteolytic cleavage site. The higher levels of reporter expression using this strategy resulted in efficient visualization of rSARS-CoV-2 in infected cultured cells and excised lungs or whole organism of infected K18 human angiotensin converting enzyme 2 (hACE2) transgenic mice. Importantly, real-time viral infection was readily tracked using a noninvasive in vivo imaging system and allowed us to rapidly identify antibodies which are able to neutralize SARS-CoV-2 infection in vivo. Notably, these reporter-expressing rSARS-CoV-2, in which a viral gene was not deleted, not only retained wild-type (WT) virus-like pathogenicity in vivo but also exhibited high stability in vitro and in vivo, supporting their use to investigate viral infection, dissemination, pathogenesis, and therapeutic interventions for the treatment of SARS-CoV-2 in vivo.

 

See: https://www.pnas.org/content/118/41/e2111593118

 

Fig. 1.

Generation of an rSARS-CoV-2 expressing Venus-2A. (A) Schematic representation of the BAC for generation of rSARS-CoV-2/Venus-2A. The sequence encoding the fusion construct Venus-2A was inserted in the viral genome of SARS-CoV-2 in the BAC. The viral ORF8, the intergenic sequences between ORF8 and N (ACAAACTAAA), Venus (green), the PTV-1 2A (light blue), and the viral N (dark blue) are indicated. (B) Vero E6 cells were mock infected or infected with rBAC-SARS-CoV-2/WT or rSARS-CoV-2/Venus-2A for 48 h, fixed, and immunostained with a MAb against the viral N protein (1C7C7). Cell nuclei were stained with DAPI. Representative images are shown. (Scale bars, 100 μm.) (C) Whole cell lysates from Vero E6 cells mock infected or infected with rSARS-CoV-2 WT or Venus-2A for 48 h were subjected to Western blot analysis using antibodies against Venus and the viral N protein (1C7C7). β-actin was used as a loading control. (D) Total cellular RNA from Vero E6 cells mock infected or infected with WT or Venus-2A rSARS-CoV-2 was isolated at 48 hpi. RT-PCR was used to amplify Venus (Top) or the region between the ORF8 and N proteins (Bottom), and the products were separated on a 0.7% agarose gel.

Back      Print      View: 123

[ Other News ]___________________________________________________
  • Genome-wide analysis of autophagy-associated genes in foxtail millet (Setaria italica L.) and characterization of the function of SiATG8a in conferring tolerance to nitrogen starvation in rice.
  • Arabidopsis small nucleolar RNA monitors the efficient pre-rRNA processing during ribosome biogenesis
  • XA21-specific induction of stress-related genes following Xanthomonas infection of detached rice leaves.
  • Reducing the Use of Pesticides with Site-Specific Application: The Chemical Control of Rhizoctonia solani as a Case of Study for the Management of Soil-Borne Diseases
  • OsJRL, a rice jacalin-related mannose-binding lectin gene, enhances Escherichia coli viability under high-salinity stress and improves salinity tolerance of rice.
  • Production of lipopeptide biosurfactants by Bacillus atrophaeus 5-2a and their potential use in microbial enhanced oil recovery.
  • GhABF2, a bZIP transcription factor, confers drought and salinity tolerance in cotton (Gossypium hirsutum L.).
  • Resilience of cassava (Manihot esculenta Crantz) to salinity: implications for food security in low-lying regions.
  • Cellulose synthase complexes act in a concerted fashion to synthesize highly aggregated cellulose in secondary cell walls of plants
  • No adverse effects of transgenic maize on population dynamics of endophytic Bacillus subtilis strain B916-gfp
  • Identification and expression analysis of OsLPR family revealed the potential roles of OsLPR3 and 5 in maintaining phosphate homeostasis in rice
  • Functional analysis of molecular interactions in synthetic auxin response circuits
  • Titanium dioxide nanoparticles strongly impact soil microbial function by affecting archaeal nitrifiers.
  • Inducible Expression of the De-Novo Designed Antimicrobial Peptide SP1-1 in Tomato Confers Resistance to Xanthomonas campestris pv. vesicatoria.
  • Toward combined delignification and saccharification of wheat straw by a laccase-containing designer cellulosome
  • SNP-based discovery of salinity-tolerant QTLs in a bi-parental population of rice (Oryza sativa)
  • Pinpointing genes underlying the quantitative trait loci for root-knot nematode resistance in palaeopolyploid soybean by whole genome resequencing.
  • Transcriptome- Assisted Label-Free Quantitative Proteomics Analysis Reveals Novel Insights into Piper nigrum -Phytophthora capsici Phytopathosystem.
  • Brassinosteroids participate in the control of basal and acquired freezing tolerance of plants
  • Rapid hyperosmotic-induced Ca2+ responses in Arabidopsis thaliana exhibit sensory potentiation and involvement of plastidial KEA transporters

 

Designed & Powered by WEBSO CO.,LTD