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Functional analysis of the 5′ flanking sequence of a vaccinia virus late gene.
Author(s) -
Bertholet C.,
Stocco P.,
Van Meir E.,
Wittek R.
Publication year - 1986
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1002/j.1460-2075.1986.tb04449.x
Subject(s) - biology , 5' flanking region , gene , virology , genetics , vaccinia , sequence (biology) , virus , sequence analysis , gene expression , recombinant dna , promoter
A series of mutations, including 5′ and 3′ deletions, as well as insertions were introduced into the 5′ flanking nucleotide sequence of a vaccinia virus late gene. This DNA has been shown previously to contain all the necessary elements for correct regulation of the gene most probably transcribed by the viral RNA polymerase. To facilitate the assays, the mutated DNA was fused to the chloramphenicol acetyltransferase gene and inserted into the genome of live vaccinia virus. The effects of the mutations on expression of the chimeric gene were studied by both enzyme assays and nuclease S1 analysis. The results showed that 5′ deletions up to about 15 bp from the putative initiation site of transcription still yielded high levels of gene expression. All mutations, however, that deleted the authentic late mRNA start site, abolished promoter activity.