A toolkit for high-throughput, cross-species gene engineering in Drosophila

We generated two complementary genomic fosmid libraries for Drosophila melanogaster and Drosophila pseudoobscura that permit seamless modification of large genomic clones by high-throughput recombineering and direct transgenesis. The fosmid transgenes recapitulated endogenous gene expression patterns. These libraries, in combination with recombineering technology, will be useful to rescue mutant phenotypes, allow imaging of gene products in living flies and enable systematic analysis and manipulation of gene activity across species.

[1]  Hugo J. Bellen,et al.  P[acman]: A BAC Transgenic Platform for Targeted Insertion of Large DNA Fragments in D. melanogaster , 2006, Science.

[2]  R. Maeda,et al.  An optimized transgenesis system for Drosophila using germ-line-specific φC31 integrases , 2007, Proceedings of the National Academy of Sciences.

[3]  M. Ashburner,et al.  Systematic determination of patterns of gene expression during Drosophila embryogenesis , 2002, Genome Biology.

[4]  N. Perrimon,et al.  Exploiting position effects and the gypsy retrovirus insulator to engineer precisely expressed transgenes , 2008, Nature Genetics.

[5]  Frank Buchholz,et al.  A new logic for DNA engineering using recombination in Escherichia coli , 1998, Nature Genetics.

[6]  G. Rubin,et al.  Global analysis of patterns of gene expression during Drosophila embryogenesis , 2007, Genome Biology.

[7]  E. Wimmer,et al.  A versatile vector set for animal transgenesis , 2000, Development Genes and Evolution.

[8]  A. Hyman,et al.  Building a spindle of the correct length in human cells requires the interaction between TPX2 and Aurora A , 2008, The Journal of cell biology.

[9]  A. Hyman,et al.  A recombineering pipeline for functional genomics applied to Caenorhabditis elegans , 2006, Nature Methods.

[10]  F. Del Bene,et al.  Optical Sectioning Deep Inside Live Embryos by Selective Plane Illumination Microscopy , 2004, Science.

[11]  Michele P Calos,et al.  Construction of transgenic Drosophila by using the site-specific integrase from phage phiC31. , 2004, Genetics.

[12]  Karl Mechtler,et al.  BAC TransgeneOmics: a high-throughput method for exploration of protein function in mammals , 2008, Nature Methods.

[13]  Bassem A. Hassan,et al.  Recombineering-mediated tagging of Drosophila genomic constructs for in vivo localization and acute protein inactivation , 2008, Nucleic acids research.

[14]  Torsten Rohlfing,et al.  Bead-based mosaicing of single plane illumination microscopy images using geometric local descriptor matching , 2009, Medical Imaging.

[15]  M. Klingler,et al.  Genetic techniques: A universal marker for transgenic insects , 1999, Nature.