We have adapted a bacterial CRISPR RNA/Cas9 system to precisely engineer the Drosophila genome and report that Cas9-mediated genomic modifications are efficiently transmitted through the germline. This RNA-guided Cas9 system can be rapidly programmed to generate targeted alleles for probing gene function in Drosophila.

Axons and dendrites differ in both microtubule (MT) organization and in the organelles and proteins they contain. Here we show that the MT motor dynein plays a critical role in polarized transport and in controlling the orientation of axonal MTs in fly dendritic arborisation (da) neurons. Changes in organelle distribution within the dendritic arbors of dynein mutant neurons correlate with a proximal shift in dendritic branch position. Dynein is also necessary for the dendrite-specific localization of Golgi outposts and the ion channel Pickpocket. Axonal MTs are normally oriented uniformly plus end-distal, but without dynein axons contain both plus and minus end-distal MTs. These data suggest that dynein is required for the distinguishing properties of the axon and dendrites: without dynein, dendritic organelles and proteins enter the axon and the axonal MTs are no longer uniform in polarity.

Multiple stringent confinement strategies should be used whenever possible

The CRISPR (clustered regularly interspaced short palindromic repeat)–Cas9 (CRISPR-associated nuclease 9) system is poised to transform developmental biology by providing a simple, efficient method to precisely manipulate the genome of virtually any developing organism. This RNA-guided nuclease (RGN)-based approach already has been effectively used to induce targeted mutations in multiple genes simultaneously, create conditional alleles, and generate endogenously tagged proteins. Illustrating the adaptability of RGNs, the genomes of >20 different plant and animal species as well as multiple cell lines and primary cells have been successfully modified. Here we review the current and potential uses of RGNs to investigate genome function during development.