Genetic Constructs: Molecular Tools for the Assembly of Environmental Bacterial Biosensors
One of the many possibilities opened by the onset of recombinant DNA technology is the construction of gene fusions between bacterial promoters and reporter genes. This allows the readout of promoter activity as a function of the presence in the medium of
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Applications: Biomonitoring
19 Genetic Constructs: Molecular Tools for the Assembly of Environmental Bacterial Biosensors A. de las Heras . V. de Lorenzo* Systems Biology Program, Centro Nacional de Biotecnologı´a-CSIC, Campus de Cantoblanco, Madrid, Spain *[email protected] 1
Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2654
2 Broad Host Range Plasmids . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2654 2.1 Promoter Probe Broad Host Range Vectors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2656 3
Construction of (Meta)Genomic Libraries with Cosmid Vectors . . . . . . . . . . . . . . . . . 2658
4 Genomic Integration of DNA Fragments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2658 4.1 Transposon-Mediated Genomic Integration of DNA Fragments . . . . . . . . . . . . . . . . . . . 2659 4.2 attB/attP Phage Integrase System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2662 5
Site-Specific Recombination Systems . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2662
6 Gene Replacement . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2664 6.1 Counterselectable Markers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2665 6.2 Allelic Exchange Methods Based on I-SceI Endonuclease Cleaving . . . . . . . . . . . . . . . . 2667 7
Selection and Counterselection Methods for Screening Libraries of Mutant Regulators . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2667
8
Implantation of Orthogonal Regulatory Systems . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2669
9
Outlook . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2669
K. N. Timmis (ed.), Handbook of Hydrocarbon and Lipid Microbiology, DOI 10.1007/978-3-540-77587-4_197, # Springer-Verlag Berlin Heidelberg, 2010
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Genetic Constructs: Molecular Tools for the Assembly of Environmental Bacterial Biosensors
Abstract: One of the many possibilities opened by the onset of recombinant DNA technology is the construction of gene fusions between bacterial promoters and reporter genes. This allows the readout of promoter activity as a function of the presence in the medium of a given physical or chemical stimulus. The availability of reporter products with optical outputs (light emission, fluorescence, enzymes for chromogenic or luminogenic substrates etc.) has permitted the design of a plethora of whole-cell biosensors for detection and even quantification of predetermined chemical species. Yet, an
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