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

Författare

Summary, in English

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.

Publiceringsår

2003

Språk

Engelska

Sidor

647-661

Publikation/Tidskrift/Serie

Genetics

Volym

163

Issue

2

Dokumenttyp

Artikel i tidskrift

Förlag

Genetics Society of America

Ämne

  • Developmental Biology

Status

Published

Forskningsgrupp

  • Invertebrate Developmental Biology, Udo Haecker's group

ISBN/ISSN/Övrigt

  • ISSN: 0016-6731