Katsuyama, T., Akmammedov, A., Seimiya, M., Hess, S.C., Sievers, C., Paro, R. (2013). An efficient strategy for TALEN-mediated genome engineering in Drosophila. Nucleic Acids Res. 41(17): e163.
FlyBase ID
FBrf0222800
Publication Type
Research paper
Abstract
In reverse genetics, a gene's function is elucidated through targeted modifications in the coding region or associated DNA cis-regulatory elements. To this purpose, recently developed customizable transcription activator-like effector nucleases (TALENs) have proven an invaluable tool, allowing introduction of double-strand breaks at predetermined sites in the genome. Here we describe a practical and efficient method for the targeted genome engineering in Drosophila. We demonstrate TALEN-mediated targeted gene integration and efficient identification of mutant flies using a traceable marker phenotype. Furthermore, we developed an easy TALEN assembly (easyT) method relying on simultaneous reactions of DNA Bae I digestion and ligation, enabling construction of complete TALENs from a monomer unit library in a single day. Taken together, our strategy with easyT and TALEN-plasmid microinjection simplifies mutant generation and enables isolation of desired mutant fly lines in the F1 generation.
