Versatile genetic paintbrushes: Brainbow technologies.
Bottom Line: While being continuously refined, Brainbow technologies have thus found a firm place in the genetic toolboxes of developmental and neurobiologists.For further resources related to this article, please visit the WIREs website.The authors have declared no conflicts of interest for this article.
Affiliation: MRC National Institute for Medical Research, Division of Molecular Neurobiology, London, UK.Show MeSH
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Mentions: Exploiting the expanding FP color palette and site-specific recombination technologies, multicolor labeling was first achieved by the Brainbow system devised by Livet et al. for mice.2 This creative approach takes advantage of the Cre-lox system to stochastically drive the expression of one of three or four FPs from a single transgene in genetically defined cell populations. Brainbow transgenes follow two principles. The Brainbow-1 strategy relies on Cre-mediated excision of DNA fragments using heterospecific lox sites (Figure 2(a)). In Brainbow-1.0 and -1.1 transgenes, three lox pairs (loxN, lox2272, and loxP) are astutely positioned in the same orientation adjacent to three or four linearly arranged FP-encoding cDNAs. These are each followed by polyA termination sequences to prevent transcriptional read-through. The FP located closest to the promoter is expressed by default. Upon Cre activation, site-specific recombination between identical lox pairs causes the excision of one, two, or three FP sequences. Consequently, new FPs are randomly positioned closest to the promoter. This leads to the stable, mutually exclusive expression of one of three or four FPs per cell in a tissue. By contrast, the Brainbow-2 strategy makes use of inversion and excision events between a single type of recombination site, loxP (Figure 2(b)). The coding sequences of two FPs are arranged in opposite orientations in an invertible cassette flanked by inward-facing loxP sites. Brainbow-2.0 contains one such cassette, and Cre-mediated inversion results in the differential expression of two markers. Brainbow-2.1 transgenes consist of two adjacent cassettes. Recombination of loxP pairs in opposite or identical orientation leads to inversion and excision of cassettes, respectively. This results in four color-outcomes. Because inversions are reversible, transient Cre expression is required. Brainbow transgenes are controlled by the Thy-1 enhancer to activate expression in neurons or glia, while recombination events are mediated by ubiquitous or tissue-specific Cre transgenes.
Affiliation: MRC National Institute for Medical Research, Division of Molecular Neurobiology, London, UK.