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Nucleic Acids Research, 1988, Vol. 16, No. 15 7269-7285
© 1988

Articles

Transcription initiation at the tet promoter and effect of mutations

Calvin B. Harley2,*,, Jonathon Lawrie3,, Mary Betlach, Roberto Crea1,4,, Herbert W. Boyer and Joe Hedgpeth5,

Howard Hughes Medical Institute and Department of Biochemistiy and Biophysics, University of California San Franciso, CA 94143-0448, USA 1Genentech, Inc. 460 Point San Bruno Boulevard, South San Francisco, CA 94080, USA

*To whom correspondence should be addressed

2Present address: Department of Biochemistry, McMaster University, Hamilton, Ontario, L8N 3Z5, Canada,

3Present address: Amoco, Five Oak Terrace, Milford, MA 01757, USA

4Present address: Creative Biomolecules, Inc., 35 South Street, Hopkinton, MA 01748, USA

5Present address: CODON, 213 East Grand Avenue, South San Francisco, CA 94080, USA

Received June 4, 1988. Accepted June 27, 1988.

we have identified the startpoint for transcription in vitro of the tetracycline resistance gene (tet) of pBR322 and several deletion and insertion mutations which alter tet promoter structure Tetracycline resistance in host bacteria correlates qualitatively with the efficiency of DNA fragments from these plasmids to promote tet tranacription in vitro. Only in active promoters could we find by computer analysis promoter structures in which the –10 and –35 sequences. and the relative spacing of the two regions agree with consensus sequence determinants. These data support the current model of the E. coli promoter sequence. Two promoter mutants gave heter 5' termini with additional A residues not encoded by the DNA sequence.


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