Zinke, Andrea, Tinney, Matthew, Hasse, Susanne, Pozniakovski, Andrei, Waterston, Robert, Rupprecht, Michaela, Janette, Judith, Angerman, Karolin, Murray, John, Schanze, Kristin, Kim, Stuart, Hyman, Anthony, Vinis, Elizabeth, Stewart, Francis, Ernst, Susanne, Reinke, Valerie, Schloissnig, Siegfried, Snyder, Michael, Teichgraber, Tina, Sarov, Mihail S.
[
International Worm Meeting,
2011]
We have developed a genome wide tag based platform for rapid in vivo function analysis of any C. elegans protein of interest. We used high-throughput recombineering to systematically engineer a "synthetic genome" collection of large genomic fragments caring fluorescent and affinity tagged alleles for each protein coding gene in the C. elegans genome. Upon genome integration these large transgenes behave like additional alleles expressing a tagged version of the protein of interest, which can be localized, biochemically purified or quantified using generic, tag based assays. Because all cis acting elements are typically included in a construct of this size the endogenous regulation mediated by regulatory proteins and RNAs at the level of transcription, splicing, message turnover and translation are maintained and can reveal fine-grained expression pattern dynamics that are not possible with the traditional cDNA based transgenes or promoter reporter constructs. The resource was successfully utilized in the modENCODE project to describe transcription factor localization at both molecular level (by ChIP-seq) and cellular level (by in vivo fluorescent microscopy).
The availability of the TransgeneOme resource opens the possibility to uncover the function of thousands of previously unstudied proteins, and would eventually allow us to build a systems wide atlas of protein localizations and interactions within the context of a multicellular model organism. Distribution of the effort throughout the community can significantly cut down the time required to achieve this goal. The use of a standard tag based techniques would make independently obtained data more comparable and easily reproducible. Finally, many of the transgenes would prove useful as reporters for loss of function studies, which will dramatically expand the accessible phenotypic space for both classic and reverse genetic approaches.