1887

Abstract

The regulatory elements which control the processes of virus replication and gene expression in the genus are uncharacterized in terms of both their locations within genome segments and their specific functions. The reverse genetics system for the type species, , has been used in combination with RNA secondary structure prediction to identify and map the positions of -acting regions within genome segment 10. Through the simultaneous introduction of variability at multiple nucleotide positions in the rescue RNA population, the functional contribution of these positions was used to map regions containing -acting elements essential for virus viability. Nucleotides that were individually lethal when varied mapped within a region of predicted secondary structure involving base pairing between the 5′ and 3′ ends of the transcript. An extended region of predicted perfect base pairing located within the 3′ untranslated region of the genome segment was also found to be required for virus viability. In contrast to the identification of individually lethal mutations, gross alteration of the composition of this predicted stem region was possible, providing the base-pairing potential between the two strands was maintained, identifying a structural feature predicted to be conserved throughout the genus. The approach of identifying -acting sequences through sequencing the recovered virus following the rescue of a degenerate RNA population is broadly applicable to viruses where reverse genetics is available.

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2015-10-01
2019-10-14
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