Abstract
With the availability of complete genome sequence for Drosophila melanogaster, one of the next strategic goals for fly researchers is a complete gene knockout collection. The P-element transposon1, the workhorse of D. melanogaster molecular genetics, has a pronounced nonrandom insertion spectrum2. It has been estimated that 87% saturation of the ∼13,500-gene complement of D. melanogaster3 might require generating and analyzing up to 150,000 insertions2. We describe specific improvements to the lepidopteran transposon piggyBac4 and the P element that enabled us to tag and disrupt genes in D. melanogaster more efficiently. We generated over 29,000 inserts resulting in 53% gene saturation and a more diverse collection of phenotypically stronger insertional alleles. We found that piggyBac has distinct global and local gene-tagging behavior from that of P elements. Notably, piggyBac excisions from the germ line are nearly always precise, piggyBac does not share chromosomal hotspots associated with P and piggyBac is more effective at gene disruption because it lacks the P bias for insertion in 5′ regulatory sequences.
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Acknowledgements
We thank the many other people at Exelixis whose intellectual and technical contributions made this work possible, particularly our colleagues in the Genomics, Informatics and Genetics departments.
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The authors are presently or were recently employed (while engaged in research on this project) by Exelixis. They may hold shares of common stock in Exelixis.
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Thibault, S., Singer, M., Miyazaki, W. et al. A complementary transposon tool kit for Drosophila melanogaster using P and piggyBac. Nat Genet 36, 283–287 (2004). https://doi.org/10.1038/ng1314
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DOI: https://doi.org/10.1038/ng1314
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