Novel TALEN-generated mCitrine-FANCD2 fusion reporter mouse model for in vivo research of DNA damage response

• Published in: DNA repair Vol. 94; p. 102936
• Main Authors: Sabol, Maja, Akbudak, M. Aydın, Fricova, Dominika, Beck, Inken, Sedlacek, Radislav
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.10.2020
• Subjects: Animals; Bacterial Proteins; DNA Damage; DNA Repair; Fanconi anemia; Fanconi Anemia Complementation Group D2 Protein - genetics; Genes, Reporter; Luminescent Proteins; mCitrine; Mice; Mice, Inbred C57BL; Models, Animal; Mouse; Recombinant Fusion Proteins; TALEN; Transcription Activator-Like Effector Nucleases; mCitrine; Fanconi anemia; Mouse; DNA damage; TALEN
• ISSN: 1568-7864; 1568-7856
• DOI: 10.1016/j.dnarep.2020.102936

•TALENs used for correct insertion of a fluorescent tag into the native locus of a target gene in a mouse model.•In-frame insertion generates a fluorescently tagged protein without overexpression and with its physiological regulation intact.•Citrine-tagged FANCD2 localizes to the sites of DNA damage after DNA damaging agents such as irradiation or doxorubicin. Reporter gene mouse lines are routinely used for studies related to functional genomics, proteomics, cell biology or cell-based drug screenings, and represent a crucial platform for in vivo research. In the generation of knock-in reporter lines, new gene targeting methods provide several advantages over the standard transgenic techniques. First of all, specific targeting of the genome allows expression of the reporter gene under controlled conditions, whether in a specific locus in the genome or in a “safe harbor” locus. Historically, the ROSA26 locus is used for gene knock-in strategies by homologous recombination in mouse embryonic stem cells. The other preferred place for integration of the reporter transgene in the mouse genome is the endogenous promoter of a target gene. In this study, we employed TALENs to generate a reporter fusion protein expressed from its native promoter. For monitoring DNA damage response, we generated a mouse line expressing a mCitrine-tagged version of the FANCD2 protein, involved in DNA damage response and repair, and the Fanconi anemia (FA) pathway. This model could be a valuable tool for in vivo investigation of DNA damage.