Development of β-Lactamase as a Tool for Monitoring Conditional Gene Expression by a Tetracycline-Riboswitch in Methanosarcina acetivorans
Joint Authors
Rother, Michael
Geist, Miriam M.
Suess, Beatrix
Weigand, Julia E.
Demolli, Shemsi
Matschiavelli, Nicole
Source
Issue
Vol. 2014, Issue 2014 (31 Dec. 2014), pp.1-10, 10 p.
Publisher
Hindawi Publishing Corporation
Publication Date
2014-01-20
Country of Publication
Egypt
No. of Pages
10
Main Subjects
Abstract EN
The use of reporter gene fusions to assess cellular processes such as protein targeting and regulation of transcription or translation is established technology in archaeal, bacterial, and eukaryal genetics.
Fluorescent proteins or enzymes resulting in chromogenic substrate turnover, like β-galactosidase, have been particularly useful for microscopic and screening purposes.
However, application of such methodology is of limited use for strictly anaerobic organisms due to the requirement of molecular oxygen for chromophore formation or color development.
We have developed β-lactamase from Escherichia coli (encoded by bla) in conjunction with the chromogenic substrate nitrocefin into a reporter system usable under anaerobic conditions for the methanogenic archaeon Methanosarcina acetivorans.
By using a signal peptide of a putative flagellin from M.
acetivorans and different catabolic promoters, we could demonstrate growth substrate-dependent secretion of β-lactamase, facilitating its use in colony screening on agar plates.
Furthermore, a series of fusions comprised of a constitutive promoter and sequences encoding variants of the synthetic tetracycline-responsive riboswitch (tc-RS) was created to characterize its influence on translation initiation in M.
acetivorans.
One tc-RS variant resulted in more than 11-fold tetracycline-dependent regulation of bla expression, which is in the range of regulation by naturally occurring riboswitches.
Thus, tc-RS fusions represent the first solely cis-active, that is, factor-independent system for controlled gene expression in Archaea.
American Psychological Association (APA)
Demolli, Shemsi& Geist, Miriam M.& Weigand, Julia E.& Matschiavelli, Nicole& Suess, Beatrix& Rother, Michael. 2014. Development of β-Lactamase as a Tool for Monitoring Conditional Gene Expression by a Tetracycline-Riboswitch in Methanosarcina acetivorans. Archaea،Vol. 2014, no. 2014, pp.1-10.
https://search.emarefa.net/detail/BIM-493667
Modern Language Association (MLA)
Demolli, Shemsi…[et al.]. Development of β-Lactamase as a Tool for Monitoring Conditional Gene Expression by a Tetracycline-Riboswitch in Methanosarcina acetivorans. Archaea No. 2014 (2014), pp.1-10.
https://search.emarefa.net/detail/BIM-493667
American Medical Association (AMA)
Demolli, Shemsi& Geist, Miriam M.& Weigand, Julia E.& Matschiavelli, Nicole& Suess, Beatrix& Rother, Michael. Development of β-Lactamase as a Tool for Monitoring Conditional Gene Expression by a Tetracycline-Riboswitch in Methanosarcina acetivorans. Archaea. 2014. Vol. 2014, no. 2014, pp.1-10.
https://search.emarefa.net/detail/BIM-493667
Data Type
Journal Articles
Language
English
Notes
Includes bibliographical references
Record ID
BIM-493667