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Amplification of maize ribulose bisphosphate carboxylase large subunit synthesis in E. coli by transcriptional fusion with the lambda N operon

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Summary

The maize chloroplast gene coding for the large subunit of ribulose bisphosphate carboxylase { 3-phospho-D-glycerate carboxy-lyase (dimerizing), EC 4.1.1.39} has been placed under the transcriptional control of the bacteriophage lambda promoter PL, by fusion with the lambda N operon located on a multicopy plasmid. Transcription from PL was repressed at 32° C by the presence in the E. coli chromosome of a cIts gene that specifies a temperature-sensitive repressor. After inactivation of the repressor at 45°C unmoderated transcription of the chloroplast gene occurred from the PL promoter. Translation was probably initiated from a chloroplast Shine-Dalgarno sequence located five nucleotides from the N-terminal methionine initiation codon to yield a polypeptide the same size as that synthesised in maize. This direct translation results in a level of expression of the chloroplast gene corresponding to approximately 2% of the total E. coli cell protein as ribulose bisphosphate carboxylase large subunits. Transcriptional fusions with the lambda N operon should provide a generally applicable, simple method for the amplification and regulation of chloroplast gene expression in E. coli.

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  1. Plant Breeding Institute, Trumpington, CB2 2LQ, Cambridge, UK

    A. A. Gatenby & J. A. Castleton

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  1. A. A. Gatenby
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  2. J. A. Castleton
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Communicated by J. Schell

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Gatenby, A.A., Castleton, J.A. Amplification of maize ribulose bisphosphate carboxylase large subunit synthesis in E. coli by transcriptional fusion with the lambda N operon. Mol Gen Genet 185, 424–429 (1982). https://doi.org/10.1007/BF00334134

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  • DOI: https://doi.org/10.1007/BF00334134

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