High Throughput Protein Trapping in Drosophila. M.H. Buszczak ¹, X. Morin ², A.T. Quinones ³, W. Chia ², L. Cooley ^3,4. 1) Dept MCDB, Yale Univ, New Haven, CT; 2) Institute of Molecular and Cell Biology, King's College London, Guy's Hospital, London, UK; 3) Dept Genetics, Yale Univ, New Haven, CT; 4) Dept Cell Biology, Yale Univ, New Haven, CT.

   Exon trapping, also referred to as protein trapping, reveals information on both the expression pattern and sub-cellular localization of a given protein. This technique has been used with great success in mouse and Xenopus and we adapted this approach for use in Drosophila (Morin et al., in press). The sequence for the Enhanced Green Fluorescent Protein (EGFP) was used to create an "exon" that lacks a start codon and is flanked on either side by splice acceptor and donor sites. This exon has been mobilized in the Drosophila genome by placing it in a P-element. We have started to perform a high throughput screen for new EGFP positive inserts by using a COPAS Drosophila embryo sorter. Hundreds of new inserts have been isolated and characterization of these EGFP positive lines reveals an incredibly diverse array of protein expression and sub-cellular localization patterns. Trapped genes are now being cloned using a combination of inverse PCR and RT-PCR. Initial results suggest that, in most cases, fusing EGFP to endogenous proteins does not affect the fidelity of protein localization. We have begun to analyze the ovarian expression patterns of trapped genes. This analysis reveals EGFP fusion proteins localize to ring canals, actin bundles, sub-nuclear domains, the posterior and anterior of the oocyte and previously uncharacterized structures in both follicle cells and nurse cells. Protein trapping is an efficient way of identifying proteins that localize to specific cell types and sub-cellular structures. The technique also contributes to the goal of producing a transposon insertion allele in every Drosophila open reading frame.