Beyond toothpicks: new methods for isolating mutant bacteria

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Abstract
New single-cell screening technologies have revolutionized research into the genetics, evolution and biotechnological applications of bacteria. This Review focuses on the uses of autofluorescent proteins (AFPs) and fluorescence techniques such as Förster resonance energy transfer (FRET) by microbiologists. Flow cytometry, which is a staple tool of eukaryotic cell biologists, is gaining popularity among microbiologists and the use of this technique is described here. The applications of microfluidic cell sorters and microfluidics, which enable mutant cells to be examined in isolation rather than as a population, in microbiology are described. The application of AFP transcriptional fusions to probe the interactions between hosts and microorganisms is also discussed. The suitability of AFPs for studying the subcellular localization of proteins is described. The identification and characterization of protein–protein interactions using AFPs is discussed. The use of water–oil emulsions in enzymatic screening are highlighted. Fluorescent screening for ligand-binding proteins is a useful technology that can be scaled up for high throughput.