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Related Experiment Video

Updated: Jul 29, 2025

An Efficient Method for Adenovirus Production
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Generation and Characterization of a Replication-Competent Human Adenovirus Type 55 Encoding EGFP.

Wei Li1, Yuehong Chen1, Ye Feng1

  • 1State Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, China.

Viruses
|May 27, 2023
PubMed
Summary

A new infectious Human adenovirus 55 (HAdV-55) virus, rAdv55-dE3-EGFP, was created for rapid antiviral screening. This tool aids in quantifying neutralizing antibodies and developing new treatments for acute respiratory disease (ARD).

Keywords:
enhanced GFPhomologous recombinationhuman adenovirus type 55infectious clonereplication-competentreporter virus

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Area of Science:

  • Virology
  • Immunology
  • Molecular Biology

Background:

  • Human adenovirus 55 (HAdV-55) outbreaks cause significant acute respiratory disease (ARD) in civilian and military populations.
  • Developing antiviral inhibitors and quantifying neutralizing antibodies necessitates rapid viral infection monitoring systems.
  • Infectious virus production from plasmids is crucial for creating experimental monitoring systems.

Purpose of the Study:

  • To construct a full-length infectious cDNA clone of HAdV-55 for generating a recombinant virus.
  • To develop a rapid, high-throughput assay for HAdV-55 neutralization testing and antiviral screening.

Main Methods:

  • Bacteria-mediated recombination was used to create the pAd55-FL clone of the HAdV-55 genome.
  • A green fluorescent protein (GFP) expression cassette replaced the E3 region, creating pAd55-dE3-EGFP.
  • The recombinant virus rAdv55-dE3-EGFP was rescued and characterized for stability and replication.

Main Results:

  • The recombinant virus rAdv55-dE3-EGFP is genetically stable and replicates similarly to wild-type HAdV-55 in cell culture.
  • rAdv55-dE3-EGFP effectively quantifies neutralizing antibodies in sera, aligning with traditional microneutralization assays.
  • The assay demonstrated utility in antiviral screening using A549 cells.

Conclusions:

  • The rAdv55-dE3-EGFP virus is a reliable tool for rapid HAdV-55 neutralization testing.
  • This high-throughput assay facilitates efficient antiviral screening against HAdV-55.
  • The developed system supports the development of countermeasures against HAdV-55-induced ARD.