A piggyBac transposon gene trap for the analysis of gene expression and function in Drosophila

Genetics
Christopher P Bonin, Richard S Mann

Abstract

P-element-based gene and enhancer trap strategies have provided a wealth of information on the expression and function of genes in Drosophila melanogaster. Here we present a new vector that utilizes the simple insertion requirements of the piggyBac transposon, coupled to a splice acceptor (SA) site fused to the sequence encoding enhanced green fluorescent protein (EGFP) and a transcriptional terminator. Mobilization of the piggyBac splice site gene trap vector (PBss) was accomplished by heat-shock-induced expression of piggyBac transposase (PBase). We show that insertion of PBss into genes leads to fusions between the gene's mRNA and the PBss-encoded EGFP transcripts. As heterozygotes, these fusions report the normal pattern of expression of the trapped gene. As homozygotes, these fusions can inactivate the gene and lead to lethality. Molecular characterization of PBss insertion events shows that they are single copy, that they always occur at TTAA sequences, and that splicing utilizes the engineered splice site in PBss. In those instances where protein-EGFP fusions are predicted to occur, the subcellular localization of the wild-type protein can be inferred from the localization of the EGFP fusion protein. These experiments hi...Continue Reading

References

Oct 5, 1990·Journal of Molecular Biology·S F AltschulD J Lipman
Dec 1, 1987·Proceedings of the National Academy of Sciences of the United States of America·C J O'Kane, W J Gehring
Nov 21, 1995·Proceedings of the National Academy of Sciences of the United States of America·A C SpradlingG M Rubin
Jul 3, 1995·Proceedings of the National Academy of Sciences of the United States of America·W C SkarnesR S Beddington
Feb 11, 1994·Science·M ChalfieD C Prasher
Sep 1, 1996·Insect Biochemistry and Molecular Biology·D A O'Brochta, P W Atkinson
Jan 15, 2000·Insect Molecular Biology·A M Handler, R A Harrell
Feb 7, 2001·Genetics·T LukacsovichD Yamamoto
Feb 15, 2001·Nature Biotechnology·E E FurlongM P Scott
Mar 10, 2001·Nature·P A LeightonM Tessier-Lavigne
Dec 14, 2001·Proceedings of the National Academy of Sciences of the United States of America·X MorinW Chia

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Citations

Jun 19, 2007·Nature Genetics·Sen WuMario R Capecchi
Jun 27, 2008·Proceedings of the National Academy of Sciences of the United States of America·Wei WangPentao Liu
Jan 26, 2010·BMC Developmental Biology·Donald A YergeauPaul E Mead
Dec 6, 2007·Genome Biology·Karl J ClarkScott C Fahrenkrug
Jul 19, 2013·PloS One·Azhahianambi PalavesamDavid A O'Brochta
Oct 12, 2013·PloS One·Maura A LaneMustafa K Khokha
Dec 21, 2006·Genetics·Ana T Quiñones-CoelloLynn Cooley
Mar 2, 2005·Nature Reviews. Genetics·Koen J T Venken, Hugo J Bellen
Oct 25, 2008·Archives of Insect Biochemistry and Physiology·Tamas LukacsovichDaisuke Yamamoto
Aug 19, 2011·The FEBS Journal·Claudia LeoniLuigi R Ceci
May 16, 2012·Proceedings of the National Academy of Sciences of the United States of America·Joni MacdonaldMichael J McGrew
Aug 24, 2018·G3 : Genes - Genomes - Genetics·William ReidDavid A O'Brochta
Jun 25, 2015·Microbiology Spectrum·Kosuke Yusa
Mar 5, 2014·Cold Spring Harbor Protocols·Karen M MannMichael B Mann

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