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Methods Mol Biol. 2020;2099:99-106. doi: 10.1007/978-1-0716-0211-9_8.

Quantification of the Middle East Respiratory Syndrome-Coronavirus RNA in Tissues by Quantitative Real-Time RT-PCR.

Author information

1
Departments of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA.
2
Department of Medical Laboratories Technology, College of Applied Medical Sciences, Jazan University, Jazan, Saudi Arabia.
3
Department of Medical Microbiology and Parasitology, Faculty of Medicine, King Abdulaziz University, Jeddah, Saudi Arabia. amhashem@kau.edu.sa.
4
Vaccines and Immunotherapy Unit, King Fahd Medical Research Center, King Abdulaziz University, Jeddah, Saudi Arabia. amhashem@kau.edu.sa.
5
Departments of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA. sktseng@utmb.edu.
6
Center for Biodefense and Emerging Infectious Disease, University of Texas Medical Branch, Galveston, TX, USA. sktseng@utmb.edu.

Abstract

Since the emergence of the Middle East respiratory syndrome-coronavirus (MERS-CoV) in 2012, more than 2280 confirmed human infections and 800 associated deaths had been reported to the World Health Organization. MERS-CoV is a single-stranded RNA virus that belongs to the Coronaviridae family. MERS-CoV infection leads to a variety of clinical outcomes in humans ranging from asymptomatic and mild infection to severe acute lung injury and multi-organ failure and death. To study the pathogenesis of MERS-CoV infection and development of medical countermeasures (MCMs) for MERS, a number of genetically modified mouse models have been developed, including various versions of transgenic mice expressing the human DPP4 viral receptor. Tracking and quantifying viral infection, among others, in permissive hosts is a key endpoint for studying MERS pathogenesis and evaluating the efficacy of selected MCMs developed for MERS. In addition to quantifying infectious progeny virus which requires high-containment biosafety level (BSL)-3 laboratory, here we outlined an established real-time quantitative RT-PCR (RT-qPCR)-based procedure to unequivocally quantify MERS-CoV-specific RNAs within the lungs of infected human DPP4 (hDPP4, transgenic (hDPP4 Tg) mice under a standard BSL-2 laboratory.

KEYWORDS:

Animal models; MERS-CoV; RT-qPCR

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