title: Exploring strategies for protein trapping in Drosophila.
abstract: The use of fluorescent protein tags has had a huge impact on cell biological studies in virtually every experimental system. Incorporation of coding sequence for fluorescent proteins such as green fluorescent protein (GFP) into genes at their endogenous chromosomal position is especially useful for generating GFP-fusion proteins that provide accurate cellular and subcellular expression data. We tested modifications of a transposon-based protein trap screening procedure in Drosophila to optimize the rate of recovering useful protein traps and their analysis. Transposons carrying the GFP-coding sequence flanked by splice acceptor and donor sequences were mobilized, and new insertions that resulted in production of GFP were captured using an automated embryo sorter. Individual stocks were established, GFP expression was analyzed during oogenesis, and insertion sites were determined by sequencing genomic DNA flanking the insertions. The resulting collection includes lines with protein traps in which GFP was spliced into mRNAs and embedded within endogenous proteins or enhancer traps in which GFP expression depended on splicing into transposon-derived RNA. We report a total of 335 genes associated with protein or enhancer traps and a web-accessible database for viewing molecular information and expression data for these genes.
authors.affiliation: Department of Genetics, Cellular and Developmental Biology, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
authors.forename: Ana T
authors.initials: AT
authors.lastname: Quiñones-Coello
authors.forename: Lisa N
authors.initials: LN
authors.lastname: Petrella
authors.forename: Kathleen
authors.initials: K
authors.lastname: Ayers
authors.forename: Anthony
authors.initials: A
authors.lastname: Melillo
authors.forename: Stacy
authors.initials: S
authors.lastname: Mazzalupo
authors.forename: Andrew M
authors.initials: AM
authors.lastname: Hudson
authors.forename: Shu
authors.initials: S
authors.lastname: Wang
authors.forename: Claudia
authors.initials: C
authors.lastname: Castiblanco
authors.forename: Michael
authors.initials: M
authors.lastname: Buszczak
authors.forename: Roger A
authors.initials: RA
authors.lastname: Hoskins
authors.forename: Lynn
authors.initials: L
authors.lastname: Cooley
mesh_terms: D000818:Animals; D015153:Blotting, Western; D003433:Crosses, Genetic; D017931:DNA Primers; D004251:DNA Transposable Elements / Q000235:genetics*; D030541:Databases, Genetic; D029721:Drosophila Proteins / Q000302:isolation & purification* / Q000378:metabolism; D004331:Drosophila melanogaster / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000235:genetics* / Q000378:metabolism; D016254:Mutagenesis, Insertional / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000378:metabolism; D017422:Sequence Analysis, DNA
chemical_list: D017931:DNA Primers; D004251:DNA Transposable Elements; D029721:Drosophila Proteins; D011993:Recombinant Fusion Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins