The art and design of genetic screens: Escherichia coli

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Abstract
Bacterial genetics is based on the ability to select for rare mutants among large populations (1 in >1010) on the basis of the ability of the bacteria to survive lethal challenges, grow in the absence of supplements or take advantage of different growth substrates. Bacterial genetics also uses a wide range of screens that are based on visual readouts of metabolic activity, including indicator agars (such as MacConkey and tetrazolium) and agars that contain chromogenic substrates (such as Xgal). One of the most powerful genetic systems is the lactose operon of Escherichia coli. The lac genes encode an enzyme for breaking down lactose — lacZ (β-galactosidase) — and the membrane transporter for lactose — lacY (lac permease). The lacZ gene can be fused to most other genes in many organisms. Transcriptional lacZ gene fusions produce wild-type β-galactosidase at levels that are determined by the promoter activity of the gene to which lacZ is fused. Translational lacZ gene fusions result in the production of hybrid β-galactosidase molecules that are encoded by both the gene to which lacZ is fused, and lacZ. The levels of production of the hybrid can depend on many post-transcriptional factors. LacZ fusions allow the powerful tools that have been developed for the lac system to be applied to almost any bacterial system. The properties of different LacZ fusions can provide important information about transcriptional and translational regulation as well as other cell functions, which include protein stability, protein folding, protein secretion and electron transport. Modern 'high-tech' approaches that involve genomics and microarrays can be combined with genetic analysis to yield new insights into complex biological problems such as host–pathogen interactions.

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