An efficient strategy for TALEN-mediated genome engineering in Drosophila
| Autor: | Cem Sievers, Makiko Seimiya, Tomonori Katsuyama, Arslan Akmammedov, Renato Paro, Samuel C. Hess |
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| Rok vydání: | 2013 |
| Předmět: |
Genetics
Transcription activator-like effector nuclease Endodeoxyribonucleases Mutant Genome Insect Gene targeting Computational biology Biology Genome Genome engineering Protein Structure Tertiary DNA-Binding Proteins Drosophila melanogaster Genome editing Mutagenesis Gene Targeting Coding region Animals Methods Online Genetic Engineering Homologous Recombination Gene Plasmids |
| Zdroj: | Nucleic Acids Research Nucleic Acids Research, 41 (17) |
| ISSN: | 1362-4962 0301-5610 |
| Popis: | 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 F 1 generation. Nucleic Acids Research, 41 (17) ISSN:1362-4962 ISSN:0301-5610 |
| Databáze: | OpenAIRE |
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