Studies of Coronavirus DI RNA Replication Using In Vitro Constructed DI cDNA Clones

  • Shinji Makino
  • Michael M. C. Lai
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 276)


Sequence analysis of an intracellular defective-interfering (DI) RNA, DIssE, of mouse hepatitis virus (MHV) revealed that it is composed of three noncontiguous genomic regions, representing the first 864 nucleotides of the 5′-end, an internal 748 nucleotides of the polymerase gene, and 601 nucleotides from the 3′-end of the parental MHV genome. DIssE had three base substitutions within the leader sequence and also a deletion of nine nucleotides located at the junction of the leader and the remaining genomic sequence. A system was developed for generating DI RNAs to study the mechanism of MHV RNA replication. A cDNA copy of DIssE RNA was placed downstream of T7 RNA polymerase promoter to generate DI RNAs capable of extremely efficient replication in the presence of a helper virus. We demonstrated that, in the DI RNA-transfected cells, the leader sequence of these DI RNAs was switched to that of the helper virus during one round of replication. This high-frequency leader sequence exchange was not observed if a nine-nucleotide stretch at the junction between the leader and the remaining DI sequence was deleted. This observation suggests that a free leader RNA is utilized for the replication of MHV RNA.


Leader Sequence Helper Virus Mouse Hepatitis Virus Packaging Signal Murine Coronavirus 
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Copyright information

© Plenum Press, New York 1990

Authors and Affiliations

  • Shinji Makino
    • 1
  • Michael M. C. Lai
    • 1
  1. 1.Department of MicrobiologyUniversity of Southern California, School of MedicineLos AngelesUSA

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