The Carnegie Protein Trap Library: A Versatile Tool for Drosophila Developmental Studies

Howard Hughes Medical Institute Research Laboratories, Department of Embryology, Carnegie Institution of Washington, Baltimore, Maryland 21218, USA.
Genetics (Impact Factor: 4.87). 04/2007; 175(3):1505-31. DOI: 10.1534/genetics.106.065961
Source: PubMed

ABSTRACT Metazoan physiology depends on intricate patterns of gene expression that remain poorly known. Using transposon mutagenesis in Drosophila, we constructed a library of 7404 protein trap and enhancer trap lines, the Carnegie collection, to facilitate gene expression mapping at single-cell resolution. By sequencing the genomic insertion sites, determining splicing patterns downstream of the enhanced green fluorescent protein (EGFP) exon, and analyzing expression patterns in the ovary and salivary gland, we found that 600-900 different genes are trapped in our collection. A core set of 244 lines trapped different identifiable protein isoforms, while insertions likely to act as GFP-enhancer traps were found in 256 additional genes. At least 8 novel genes were also identified. Our results demonstrate that the Carnegie collection will be useful as a discovery tool in diverse areas of cell and developmental biology and suggest new strategies for greatly increasing the coverage of the Drosophila proteome with protein trap insertions.

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Available from: Benjamin Ohlstein, Jan 14, 2014
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    • "Baz has been reported to localize at the hub-GSC interface along with E-cadherin following overexpression in the germline (nos > Baz-GFP) (Leatherman and Dinardo, 2010), which was confirmed by using independent UAS- Baz-YFP construct (see below). However, closer inspection using antibody staining and Flytrap Baz- GFP that expresses endogenous levels of Baz [CC01941 (Kelso et al., 2004; Buszczak et al., 2007)] revealed that Baz forms foci at the hub-GSC interface (referred to as the 'Baz patch' hereafter), instead of entirely colocalizing with E-cadherin (Figure 1B,C). The Baz patch is a small structure, with a size of approximately 1.5 μm, and this patch is considerably smaller than the GSC-hub interface that is marked by E-cadherin (4–6 μm) (Figure 1C). "
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    • "Although we cannot directly assess the proportion of inserts in non-essential genes that are functional, this is likely to be similar to that of essential genes. Overall, we have strong evidence for protein trap insertions in 374 annotated genes and we compared this list with the verified protein traps reported in the FlyTrap database (Morin et al., 2001; Buszczak et al., 2007; Quinones-Coello et al., 2007). Analysing the lists of trapped genes in FlyMine to account for any annotation differences, Fig. 1. "
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    • "Fly stocks useful for functional genomic studies (Table 1) can be generally grouped into (1) those in which the endogenous gene is mutated, disrupted, or tagged directly, e.g., classical alleles, deletions, Gene Disruption Project (GDP) transposon insertions, and " protein trap " lines (Morin et al. 2001; Bellen et al. 2004, 2011; Kelso et al. 2004; Buszczak et al. 2007; Cook et al. 2010a, 2012; Venken et al. 2011; Singari et al. 2014), and (2) those for which transgenesis into a neutral or unrelated location is used to introduce an exogenous reporter construct, expression driver, reagent, etc. (reviewed in Cook et al. 2010a; Perrimon et al. 2010). Some fly stocks, such as GDP MiMIC element insertions, fall into both categories, as they can be useful as mutant alleles and/or used to develop reporter constructs , protein fusions, etc. (Venken et al. 2011). "
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