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A new T7 RNA polymerase-driven expression system induced via thermoamplification of a recombinant plasmid carrying a T7 promoter-Escherichia coli lac operator.
Gene 1994; 142(1):61-6GENE

Abstract

A new temperature-regulated T7 RNA polymerase-driven transcription system has been developed. This system is based on a hybrid regulatory region: the phage T7 late promoter (PT7) linked to the Escherichia coli lac operator (Olac) [Giordano et al., Gene 84 (1989) 209-219], which was located in an earlier obtained [Mashko et al., Gene 97 (1991) 259-266] temperature-controlled amplifiable plasmid, carrying cat under the control of PT7-Olac and, in addition, lambda major early promoter-operator regions and gene cIts857. Plasmids of the pT7-Olac-cat-tsr series were stably maintained at a low-copy-number when grown at low temperature (28 degrees C). In E. coli BL21(DE3), carrying the Plac-controllable T7 RNA polymerase-encoding gene, efficient repression of cat transcription was observed, that was provided by the LacI repressor and, probably, the thermolabile repressor CIts857. At low and moderate temperatures (28/37 degrees C), this 'cooperative' repression was so tight that cat expression was not observed in the cells carrying PT7-Olac on the plasmids, even after IPTG-inducible T7 RNA polymerase biosynthesis. As a result of the thermo-amplification of the recombinant plasmids and temperature-inactivation of CIts857, expression of the T7 RNA polymerase-encoding gene was derepressed due to the titration of LacI by the increasing copies of Olac which in turn, led to the highly efficient T7 RNA polymerase-driven accumulation of CAT in the cells.

Authors+Show Affiliations

Institute of Genetics and Selection of Industrial Microorganisms, Moscow, Russia.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article

Language

eng

PubMed ID

8181758

Citation

Lebedeva, M I., et al. "A New T7 RNA Polymerase-driven Expression System Induced Via Thermoamplification of a Recombinant Plasmid Carrying a T7 promoter-Escherichia Coli Lac Operator." Gene, vol. 142, no. 1, 1994, pp. 61-6.
Lebedeva MI, Rogozhkina EV, Tsyba NA, et al. A new T7 RNA polymerase-driven expression system induced via thermoamplification of a recombinant plasmid carrying a T7 promoter-Escherichia coli lac operator. Gene. 1994;142(1):61-6.
Lebedeva, M. I., Rogozhkina, E. V., Tsyba, N. A., & Mashko, S. V. (1994). A new T7 RNA polymerase-driven expression system induced via thermoamplification of a recombinant plasmid carrying a T7 promoter-Escherichia coli lac operator. Gene, 142(1), pp. 61-6.
Lebedeva MI, et al. A New T7 RNA Polymerase-driven Expression System Induced Via Thermoamplification of a Recombinant Plasmid Carrying a T7 promoter-Escherichia Coli Lac Operator. Gene. 1994 May 3;142(1):61-6. PubMed PMID: 8181758.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - A new T7 RNA polymerase-driven expression system induced via thermoamplification of a recombinant plasmid carrying a T7 promoter-Escherichia coli lac operator. AU - Lebedeva,M I, AU - Rogozhkina,E V, AU - Tsyba,N A, AU - Mashko,S V, PY - 1994/5/3/pubmed PY - 1994/5/3/medline PY - 1994/5/3/entrez SP - 61 EP - 6 JF - Gene JO - Gene VL - 142 IS - 1 N2 - A new temperature-regulated T7 RNA polymerase-driven transcription system has been developed. This system is based on a hybrid regulatory region: the phage T7 late promoter (PT7) linked to the Escherichia coli lac operator (Olac) [Giordano et al., Gene 84 (1989) 209-219], which was located in an earlier obtained [Mashko et al., Gene 97 (1991) 259-266] temperature-controlled amplifiable plasmid, carrying cat under the control of PT7-Olac and, in addition, lambda major early promoter-operator regions and gene cIts857. Plasmids of the pT7-Olac-cat-tsr series were stably maintained at a low-copy-number when grown at low temperature (28 degrees C). In E. coli BL21(DE3), carrying the Plac-controllable T7 RNA polymerase-encoding gene, efficient repression of cat transcription was observed, that was provided by the LacI repressor and, probably, the thermolabile repressor CIts857. At low and moderate temperatures (28/37 degrees C), this 'cooperative' repression was so tight that cat expression was not observed in the cells carrying PT7-Olac on the plasmids, even after IPTG-inducible T7 RNA polymerase biosynthesis. As a result of the thermo-amplification of the recombinant plasmids and temperature-inactivation of CIts857, expression of the T7 RNA polymerase-encoding gene was derepressed due to the titration of LacI by the increasing copies of Olac which in turn, led to the highly efficient T7 RNA polymerase-driven accumulation of CAT in the cells. SN - 0378-1119 UR - https://www.unboundmedicine.com/medline/citation/8181758/A_new_T7_RNA_polymerase_driven_expression_system_induced_via_thermoamplification_of_a_recombinant_plasmid_carrying_a_T7_promoter_Escherichia_coli_lac_operator_ L2 - https://linkinghub.elsevier.com/retrieve/pii/0378-1119(94)90355-7 DB - PRIME DP - Unbound Medicine ER -