Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 1992 Jul 11;20(13):3375-81.
doi: 10.1093/nar/20.13.3375.

Homologous RNA recombination allows efficient introduction of site-specific mutations into the genome of coronavirus MHV-A59 via synthetic co-replicating RNAs

R G van der Most  1 , L Heijnen, W J Spaan, R J de Groot
Affiliations
Free PMC article

Homologous RNA recombination allows efficient introduction of site-specific mutations into the genome of coronavirus MHV-A59 via synthetic co-replicating RNAs

R G van der Most et al. Nucleic Acids Res. .
Free PMC article

Abstract

We describe a novel strategy to site-specifically mutagenize the genome of an RNA virus by exploiting homologous RNA recombination between synthetic defective interfering (DI) RNA and the viral RNA. The construction of a full-length cDNA clone, pMIDI, of a DI RNA of coronavirus MHV strain A59 was reported previously (R.G. Van der Most, P.J. Bredenbeek, and W.J.M. Spaan (1991). J. Virol. 65, 3219-3226). RNA transcribed from this construct, is replicated efficiently in MHV-infected cells. Marker mutations introduced in MIDI RNA were replaced by the wild-type residues during replication. More importantly, however, these genetic markers were introduced into viral genome: even in the absence of positive selection MHV recombinants could be isolated. This finding provides new prospects for the study of coronavirus replication using recombinant DNA techniques. As a first application, we describe the rescue of the temperature sensitive mutant MHV Albany-4 using DI-directed mutagenesis. Possibilities and limitations of this strategy are discussed.

PubMed Disclaimer

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. J Virol. 1990 Mar;64(3):1050-6 - PubMed
    1. J Gen Virol. 1986 Oct;67 ( Pt 10):2067-82 - PubMed
    1. Virology. 1990 Aug;177(2):634-45 - PubMed
    1. Virology. 1990 Aug;177(2):646-56 - PubMed
    1. J Virol. 1990 Oct;64(10):4761-6 - PubMed

Publication types