Cloning of avian tumor virus DNA fragments in plasmid pBR322: evidence for efficient transcription in E. coli from a virus-coded promoter

Ramareddy Guntaka, S. Alex Mitsialis

Research output: Contribution to journalArticle

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Abstract

Two avian tumor virus DNA fragments of 4.2 and 3.2 kb were inserted into pBR322 in the two possible orientations for each fragment. In the Escherichia coli host cells, RNA polymerase initiates transcription of large quantities (up to 0.5 to 1% of total E. coli RNA) of virus-specific RNA in the recombinant plasmids carrying the 4.2-kb fragment (pATV-6) but not in pATV-2 which contains the 3.2-kb fragment. Two SacI cleavage sites flank the putative promoter in the 4.2-kb viral insert. Deletion in the 1.2-kb SacI fragment obliterated the ability of pATV-6 to synthesize viral RNA. Digestion of the 1.2-kb SacI fragment with PvuI generates two fragments of 0.63 and 0.57 kb. Deletion of the 0.57-kb but not the 0.63-kb PvuI-SacI fragment completely eliminated the ability of the recombinant to synthesize viral RNA. These results strongly suggest that viral RNA in E. coli transcription is indeed initiated at a size present in the viral genome and that this site is localized in the 0.57-kb PvuI-SacI fragment.

Original languageEnglish (US)
Pages (from-to)113-121
Number of pages9
JournalGene
Volume12
Issue number1-2
DOIs
StatePublished - Jan 1 1980

Fingerprint

DNA Tumor Viruses
Viral RNA
Organism Cloning
Plasmids
Escherichia coli
Viruses
Viral Genome
RNA Viruses
DNA-Directed RNA Polymerases
Digestion

All Science Journal Classification (ASJC) codes

  • Medicine(all)
  • Genetics

Cite this

Cloning of avian tumor virus DNA fragments in plasmid pBR322 : evidence for efficient transcription in E. coli from a virus-coded promoter. / Guntaka, Ramareddy; Mitsialis, S. Alex.

In: Gene, Vol. 12, No. 1-2, 01.01.1980, p. 113-121.

Research output: Contribution to journalArticle

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