Tissue-specific tagging of endogenous loci inDrosophila melanogaster [article]

Kate Koles, Anna R Yeh, Avital A Rodal
2015 bioRxiv   pre-print
Fluorescent protein tags have revolutionized cell and developmental biology, and in combination with binary expression systems they enable diverse tissue-specific studies of protein function. However these binary expression systems often do not recapitulate endogenous protein expression levels, localization, binding partners, and developmental windows of gene expression. To address these limitations, we have developed a method called T-STEP (Tissue-SpecificTagging ofEndogenousProteins) that
more » ... ws endogenous loci to be tagged in a tissue specific manner. T-STEP uses a combination of efficient gene targeting and tissue-specific recombinase-mediated tag swapping to temporally and spatially label endogenous proteins. We have employed this method to GFP tag OCRL (a phosphoinositide-5-phosphatase in the endocytic pathway) and Vps35 (a Parkinson's disease-implicated component of the endosomal retromer complex) in diverse Drosophila tissues including neurons, glia, muscles, and hemocytes. Selective tagging of endogenous proteins allows for the first time cell type-specific live imaging and proteomics in complex tissues.
doi:10.1101/025635 fatcat:fkrv7evpovcrlgtvgatbr5wj3a