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PMID: 3479621 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S.

Production of infectious RNA transcripts from Sindbis virus cDNA clones: mapping of lethal mutations, rescue of a temperature-sensitive marker, and in vitro mutagenesis to generate defined mutants.

Journal of virology ·Vol. 61 ·No. 12 ·1987-12-00 ·Pages 3809-19

Rice CM, Levis R, Strauss JH, Huang HV

Abstract

We constructed full-length cDNA clones of Sindbis virus that can be transcribed in vitro by SP6 RNA polymerase to produce infectious genome-length transcripts. Viruses produced from in vitro transcripts are identical to Sindbis virus and show strain-specific phenotypes reflecting the source of RNA used for cDNA synthesis. The cDNA clones were used to confirm the mapping of the causal mutation of ts2 to the capsid protein. A general strategy for mapping Sindbis virus mutations is described and was used to identify two lethal mutations in an original full-length construct which did not produce infectious transcripts. An XbaI linker was inserted in the cDNA clone near the transcriptional start of the subgenomic mRNA; the resulting virus retains the XbaI recognition sequence, thus providing formal evidence that viruses are derived from in vitro transcripts of cDNA clones. The potential applications of the cDNA clones are discussed.

MeSH Terms
Animals Cell Line Cloning, Molecular DNA/genetics DNA, Viral/genetics Genes, Viral Genetic Markers Mutation Phenotype RNA, Messenger/genetics RNA, Viral/genetics Sindbis Virus/genetics Temperature Transcription, Genetic Transfection Virion/genetics
Chemicals
DNA, Viral Genetic Markers RNA, Messenger RNA, Viral DNA
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Rice C M
Department of Microbiology and Immunology, Washington University School of Medicine, St. Louis, Missouri 63110-1093.
Levis R
Strauss J H
Huang H V
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Article Info
Journal
Journal of virology
Abbr.
J Virol
ISSN
0022-538X
DOI
10.1128/JVI.61.12.3809-3819.1987
Published
1987-12-00
Pages
3809-19
Language
English
Region
United States
NLM ID
0113724
PMCID
PMC255997
Subset
IM
Grants
NIAID NIH HHS · AI10793 · United States
NIAID NIH HHS · AI20612 · United States
NIAID NIH HHS · AI24134 · United States
External Links
PubMed source DOI: 10.1128/JVI.61.12.3809-3819.1987 PMC Full Text View journal details
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