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piggyBac-based insertional mutagenesis and enhancer detection as a tool for functional insect genomics

Horn, C; Offen, N; Nystedt, Sverker LU ; Häcker, Udo LU and Wimmer, EA (2003) In Genetics 163(2). p.647-661
Abstract
Transposon mutagenesis provides a fundamental tool for functional genomics. Here we present a non-species-specific, combined enhancer detection and binary expression system based on the transposable element piggyBac: For the different components of this insertional mutagenesis system, we used widely applicable transposons and distinguishable broad-range transformation markers, which should enable this system to be operational in nonmodel arthropods. In a pilot screen in Drosophila melanogaster, piggyBac mutator elements on the X chromosome were mobilized in males by a Hennes-based jumpstarter element providing pigpBac transposase activity under control of the alpha1-tubulin promoter. As primary reporters in the pigoBac mutator elements, we... (More)
Transposon mutagenesis provides a fundamental tool for functional genomics. Here we present a non-species-specific, combined enhancer detection and binary expression system based on the transposable element piggyBac: For the different components of this insertional mutagenesis system, we used widely applicable transposons and distinguishable broad-range transformation markers, which should enable this system to be operational in nonmodel arthropods. In a pilot screen in Drosophila melanogaster, piggyBac mutator elements on the X chromosome were mobilized in males by a Hennes-based jumpstarter element providing pigpBac transposase activity under control of the alpha1-tubulin promoter. As primary reporters in the pigoBac mutator elements, we employed the heterologous transactivators GAL4Delta or tTA. To identify larval and adult enhancer detectors, strains carrying UASp-EYFP or TRE-EYFP as secondary reporter elements were used. Tissue-specific enhancer activities were readily observed in the GAL4Delta/UASp-based systems, but only rarely in the tTA/TRE system. Novel autosomal insertions were recovered with an average jumping rate of 80%. Of these novel insertions, 3.8% showed homozygous lethality, which was reversible by pigoBac excision. Insertions were found in both coding and noncoding regions of characterized genes and also in noncharacterized and non-P-targeted CG-number genes. This indicates that piggyBac will greatly facilitate the intended saturation mutagenesis in Drosophila. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Genetics
volume
163
issue
2
pages
647 - 661
publisher
Genetics Society of America
external identifiers
  • wos:000181417200019
  • pmid:12618403
  • scopus:0037295418
ISSN
0016-6731
language
English
LU publication?
yes
id
4395defb-fbdf-4863-b4e6-8024e4552a3d (old id 316987)
alternative location
http://www.genetics.org/cgi/content/abstract/163/2/647
date added to LUP
2007-09-13 11:12:09
date last changed
2018-06-03 03:28:51
@article{4395defb-fbdf-4863-b4e6-8024e4552a3d,
  abstract     = {Transposon mutagenesis provides a fundamental tool for functional genomics. Here we present a non-species-specific, combined enhancer detection and binary expression system based on the transposable element piggyBac: For the different components of this insertional mutagenesis system, we used widely applicable transposons and distinguishable broad-range transformation markers, which should enable this system to be operational in nonmodel arthropods. In a pilot screen in Drosophila melanogaster, piggyBac mutator elements on the X chromosome were mobilized in males by a Hennes-based jumpstarter element providing pigpBac transposase activity under control of the alpha1-tubulin promoter. As primary reporters in the pigoBac mutator elements, we employed the heterologous transactivators GAL4Delta or tTA. To identify larval and adult enhancer detectors, strains carrying UASp-EYFP or TRE-EYFP as secondary reporter elements were used. Tissue-specific enhancer activities were readily observed in the GAL4Delta/UASp-based systems, but only rarely in the tTA/TRE system. Novel autosomal insertions were recovered with an average jumping rate of 80%. Of these novel insertions, 3.8% showed homozygous lethality, which was reversible by pigoBac excision. Insertions were found in both coding and noncoding regions of characterized genes and also in noncharacterized and non-P-targeted CG-number genes. This indicates that piggyBac will greatly facilitate the intended saturation mutagenesis in Drosophila.},
  author       = {Horn, C and Offen, N and Nystedt, Sverker and Häcker, Udo and Wimmer, EA},
  issn         = {0016-6731},
  language     = {eng},
  number       = {2},
  pages        = {647--661},
  publisher    = {Genetics Society of America},
  series       = {Genetics},
  title        = {piggyBac-based insertional mutagenesis and enhancer detection as a tool for functional insect genomics},
  volume       = {163},
  year         = {2003},
}