‘Primer alignment-and-extension’: a novel mechanism of viral RNA recombination responsible for the rescue of inactivated poliovirus cDNA clones

Abstract

In the course of experiments designed to assess the potential role of alternative open reading frames (ORF) present in the 5′-terminal untranslated region (5′-UTR) of poliovirus type 1 (Mahoney strain) genomic RNA, we came across a double mutation that completely abrogated the infectivity of full-length cDNA clones. The infectivity was rescued in trans by cotransfecting COS-1 cells with short RNA transcripts of the wild-type 5′-UTR of poliovirus type 2 Lansing, provided a free 3′-OH was available. Direct sequencing of the viral RNA revealed that the infectious viruses recovered were recombinants Lansing/Mahoney, with variable points of ‘crossing-over’. A novel mechanism of RNA–RNA recombination, which we propose to call ‘primer alignment-and-extension’, is described that would explain the high rate of recombination of RNA viruses observed in natural conditions.