Rapid one-step construction of a Middle East Respiratory Syndrome (MERS-CoV) infectious clone system by homologous recombination

Aidan M Nikiforuk; Anders Leung; Bradley W M Cook; Deborah A Court; Darwyn Kobasa; Steven S Theriault

doi:10.1016/j.jviromet.2016.07.022

Rapid one-step construction of a Middle East Respiratory Syndrome (MERS-CoV) infectious clone system by homologous recombination

J Virol Methods. 2016 Oct:236:178-183. doi: 10.1016/j.jviromet.2016.07.022. Epub 2016 Jul 25.

Authors

Aidan M Nikiforuk¹, Anders Leung², Bradley W M Cook³, Deborah A Court⁴, Darwyn Kobasa⁵, Steven S Theriault⁶

Affiliations

¹ Applied Biosafety Research Program, National Microbiology Laboratory at the Canadian Science Centre for Human and Animal Health, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba, R3E 3P6, Canada; National Microbiology Laboratory at the J. C. Wilt Infectious Diseases Research Centre, Public Health Agency of Canada, 745 Logan Street, Winnipeg, Manitoba, R3E 3L5, Canada; High Containment Respiratory Viruses Group, Special Pathogens Program, National Microbiology Laboratory at the Canadian Science Centre for Human and Animal Health, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba, R3E 3R2, Canada; Department of Microbiology, The University of Manitoba, Winnipeg, Manitoba, R3T 2N2, Canada.
² High Containment Respiratory Viruses Group, Special Pathogens Program, National Microbiology Laboratory at the Canadian Science Centre for Human and Animal Health, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba, R3E 3R2, Canada.
³ Applied Biosafety Research Program, National Microbiology Laboratory at the Canadian Science Centre for Human and Animal Health, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba, R3E 3P6, Canada; National Microbiology Laboratory at the J. C. Wilt Infectious Diseases Research Centre, Public Health Agency of Canada, 745 Logan Street, Winnipeg, Manitoba, R3E 3L5, Canada; Department of Microbiology, The University of Manitoba, Winnipeg, Manitoba, R3T 2N2, Canada.
⁴ Department of Microbiology, The University of Manitoba, Winnipeg, Manitoba, R3T 2N2, Canada.
⁵ High Containment Respiratory Viruses Group, Special Pathogens Program, National Microbiology Laboratory at the Canadian Science Centre for Human and Animal Health, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba, R3E 3R2, Canada; Department of Medical Microbiology, The University of Manitoba, Winnipeg, Manitoba, R3T 2N2,Canada.
⁶ Applied Biosafety Research Program, National Microbiology Laboratory at the Canadian Science Centre for Human and Animal Health, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba, R3E 3P6, Canada; National Microbiology Laboratory at the J. C. Wilt Infectious Diseases Research Centre, Public Health Agency of Canada, 745 Logan Street, Winnipeg, Manitoba, R3E 3L5, Canada; Department of Microbiology, The University of Manitoba, Winnipeg, Manitoba, R3T 2N2, Canada. Electronic address: steven.theriault@phac-aspc.gc.ca.

Abstract

Background: Viral Infectious clone systems serve as robust platforms to study viral gene or replicative function by reverse genetics, formulate vaccines and adapt a wild type-virus to an animal host. Since the development of the first viral infectious clone system for the poliovirus, novel strategies of viral genome construction have allowed for the assembly of viral genomes across the identified viral families. However, the molecular profiles of some viruses make their genome more difficult to construct than others. Two factors that affect the difficulty of infectious clone construction are genome length and genome complexity.

Results: This work examines the available strategies for overcoming the obstacles of assembling the long and complex RNA genomes of coronaviruses and reports one-step construction of an infectious clone system for the Middle East Respiratory Syndrome coronavirus (MERS-CoV) by homologous recombination in S. cerevisiae.

Conclusions: Future use of this methodology will shorten the time between emergence of a novel viral pathogen and construction of an infectious clone system. Completion of a viral infectious clone system facilitates further study of a virus's biology, improvement of diagnostic tests, vaccine production and the screening of antiviral compounds.

Keywords: Coronavirus; Homologous recombination; Infectious clone system; Middle East Respiratory Syndrome; Molecular cloning; Reverse genetics.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Cell Line
Genetic Engineering / methods*
Homologous Recombination*
Middle East Respiratory Syndrome Coronavirus / genetics*
Middle East Respiratory Syndrome Coronavirus / physiology*
Reverse Genetics / methods*