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ToolsAli, Hanif and Murrell, J. Colin (2009) Development and validation of promoter-probe vectors for the study of methane monooxygenase gene expression in Methylococcus capsulatus Bath. Microbiology, 155 (3). pp. 761-771. ISSN 1350-0872
Full text not available from this repository. (Request a copy)Abstract
A series of integrative and versatile broad-host-range promoter-probe vectors carrying reporter genes encoding green fluorescent protein (GFP), catechol 2,3-dioxygenase (XylE) or ß-galactosidase (LacZ) were constructed for use in methanotrophs. These vectors facilitated the measurement of in vivo promoter activity in methanotrophs under defined growth conditions. They were tested by constructing transcriptional fusions between the soluble methane monooxygenase (sMMO) s54 promoter or particulate methane monooxygenase (pMMO) s70 promoter from Methylococcus capsulatus and the reporter genes. Reporter gene activity was measured under high- and low-copper growth conditions and the data obtained closely reflected transcriptional regulation of the sMMO or pMMO operon, thus demonstrating the suitability of these vectors for assessing promoter activity in methanotrophs. When ß-galactosidase expression was coupled with the fluorogenic substrate 4-methylumbelliferyl ß-d-glucuronide it yielded a sensitive and powerful screening system for detecting cells expressing this reporter gene. These data were substantiated with independent experiments using RT-PCR and RNA dot-blot analysis.
| Item Type: | Article |
|---|---|
| Faculty \ School: | Faculty of Science > School of Environmental Sciences University of East Anglia Research Groups/Centres > Theme - ClimateUEA |
| UEA Research Groups: | Faculty of Science > Research Centres > Centre for Ecology, Evolution and Conservation Faculty of Science > Research Groups > Environmental Biology |
| Depositing User: | Rhiannon Harvey |
| Date Deposited: | 27 Mar 2012 12:26 |
| Last Modified: | 20 Mar 2023 15:31 |
| URI: | https://ueaeprints.uea.ac.uk/id/eprint/38498 |
| DOI: | 10.1099/mic.0.021816-0 |
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