Captured Segment Exchange: A Strategy for Custom Engineering Large Genomic Regions in Drosophila melanogaster [Methods, Technology, and Resources]

Captured Segment Exchange: A Strategy for Custom Engineering Large Genomic Regions in Drosophila melanogaster [Methods, Technology, and Resources]:
Site-specific recombinases (SSRs) are valuable tools for manipulating genomes. In Drosophila, thousands of transgenic insertions carrying SSR recognition sites have been distributed throughout the genome by several large-scale projects. Here we describe a method with the potential to use these insertions to make custom alterations to the Drosophila genome in vivo. Specifically, by employing recombineering techniques and a dual recombinase-mediated cassette exchange strategy based on the phiC31 integrase and FLP recombinase, we show that a large genomic segment that lies between two SSR recognition-site insertions can be "captured" as a target cassette and exchanged for a sequence that was engineered in bacterial cells. We demonstrate this approach by targeting a 50-kb segment spanning the tsh gene, replacing the existing segment with corresponding recombineered sequences through simple and efficient manipulations. Given the high density of SSR recognition-site insertions in Drosophila, our method affords a straightforward and highly efficient approach to explore gene function in situ for a substantial portion of the Drosophila genome.

Long-Range Targeted Manipulation of the Drosophila Genome by Site-Specific Integration and Recombinational Resolution [Methods, Technology, and Resources]

Long-Range Targeted Manipulation of the Drosophila Genome by Site-Specific Integration and Recombinational Resolution [Methods, Technology, and Resources]:
Significant advances in genomics underscore the importance of targeted mutagenesis for gene function analysis. Here we have developed a scheme for long-range targeted manipulation of genes in the Drosophila genome. Utilizing an attP attachment site for the phiC31 integrase previously targeted to the nbs gene, we integrated an 80-kb genomic fragment at its endogenous locus to generate a tandem duplication of the region. We achieved reduction to a single copy by inducing recombination via a site-specific DNA break. We report that, despite the large size of the DNA fragment, both plasmid integration and duplication reduction can be accomplished efficiently. Importantly, the integrating genomic fragment can serve as a venue for introducing targeted modifications to the entire region. We successfully introduced a new attachment site 70 kb from the existing attP using this two-step scheme, making a new region susceptible to targeted mutagenesis. By experimenting with different placements of the future DNA break site in the integrating vector, we established a vector configuration that facilitates the recovery of desired modifications. We also show that reduction events can occur efficiently through unequal meiotic crossing over between the large duplications. Based on our results, we suggest that a collection of 1200 lines with attachment sites inserted every 140 kb throughout the genome would render all Drosophila genes amenable to targeted mutagenesis. Excitingly, all of the components involved are likely functional in other eukaryotes, making our scheme for long-range targeted manipulation readily applicable to other systems.

Constraint on ftz evolution [Evolution]

Constraint on ftz evolution [Evolution]: Despite enormous body plan variation, genes regulating embryonic development are highly conserved. Here, we probe the mechanisms that predispose ancient regulatory genes to reutilization and diversification rather than evolutionary loss. The Hox gene fushi tarazu (ftz) arose as a homeotic gene but functions as a pair-rule segmentation gene in Drosophila....

Transferring a synthetic gene circuit from yeast to mammalian cells

Transferring a synthetic gene circuit from yeast to mammalian cells:
Transferring a synthetic gene circuit from yeast to mammalian cells

Nature Communications 4, 1451 (2013). doi:10.1038/ncomms2471

Authors: Dmitry Nevozhay, Tomasz Zal & Gábor Balázsi

Developmental evidence for serial homology of the vertebrate jaw and gill arch skeleton

Developmental evidence for serial homology of the vertebrate jaw and gill arch skeleton:
Developmental evidence for serial homology of the vertebrate jaw and gill arch skeleton

Nature Communications 4, 1436 (2013). doi:10.1038/ncomms2429

Authors: J. Andrew Gillis, Melinda S. Modrell & Clare V. H. Baker

Yeast Adapts to a Changing Stressful Environment by Evolving Cross-Protection and Anticipatory Gene Regulation

Yeast Adapts to a Changing Stressful Environment by Evolving Cross-Protection and Anticipatory Gene Regulation:
Organisms can protect themselves against future environmental change. An example is cross-protection, where physiological adaptation against a present environmental stressor can protect an organism against a future stressor. Another is anticipation, where an organism uses information about its present environment to trigger gene expression and other physiological changes adaptive in future environments. "Predictive" abilities like this exist in organisms that have been exposed to periodic changes in environments. It is unknown how readily they can evolve. To answer this question, we carried out laboratory evolution experiments in the yeast Saccharomyces cerevisiae. Specifically, we exposed three replicate populations of yeast to environments that varied cyclically between two stressors, salt stress and oxidative stress, every 10 generations, for a total of 300 generations. We evolved six replicate control populations in only one of these stressors for the same amount of time. We analyzed fitness changes and genome-scale expression changes in all these evolved populations. Our populations evolved asymmetric cross protection, where oxidative stress protects against salt stress but not vice versa. Gene expression data also suggest the evolution of anticipation and basal gene expression changes that occur uniquely in cyclic environments. Our study shows that highly complex physiological states that are adaptive in future environments can evolve on very short evolutionary time scales.

Multi-channel acoustic recording and automated analysis of Drosophila courtship songs

Multi-channel acoustic recording and automated analysis of Drosophila courtship songs: Background:
Drosophila melanogaster has served as a powerful model system for genetic studies of courtship songs. To accelerate research on the genetic and neural mechanisms underlying courtship song, we have developed a sensitive recording system to simultaneously capture the acoustic signals from 32 separate pairs of courting flies as well as software for automated segmentation of songs.
Results:
Our novel hardware design enables recording of low amplitude sounds in most laboratory environments. We demonstrate the power of this system by collecting, segmenting and analyzing over 18 hours of courtship song from 75 males from five wild-type strains of Drosophila melanogaster. Our analysis reveals previously undetected modulation of courtship song features and extensive natural genetic variation for most components of courtship song. Despite having a large dataset with sufficient power to detect subtle modulations of song, we were unable to identify previously reported periodic rhythms in the inter-pulse interval of song. We provide detailed instructions for assembling the hardware and for using our open-source segmentation software.
Conclusions:
Analysis of a large dataset of acoustic signals from Drosophila melanogaster provides novel insight into the structure and dynamics of species-specific courtship songs. Our new system for recording and analyzing fly acoustic signals should therefore greatly accelerate future studies of the genetics, neurobiology and evolution of courtship song.