Constitutive and Inducible Systems for Genetic In Vivo Modification of Mouse Hepatocytes Using Hydrodynamic Tail Vein Injection
| Autor: | Roland M. Schmid, C Lechler, B Kohnke-Ertel, Ursula Ehmer, Eric K. Hubner, Thomas Rosner |
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| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
General Immunology and Microbiology General Chemical Engineering General Neuroscience Regeneration (biology) Transgene Transfection Biology Gene mutation General Biochemistry Genetics and Molecular Biology Cell biology 03 medical and health sciences 030104 developmental biology In vivo Gene expression Gene silencing Gene |
| Zdroj: | Journal of Visualized Experiments. |
| ISSN: | 1940-087X |
| DOI: | 10.3791/56613 |
| Popis: | In research models of liver cancer, regeneration, inflammation, and fibrosis, flexible systems for in vivo gene expression and silencing are highly useful. Hydrodynamic tail vein injection of transposon-based constructs is an efficient method for genetic manipulation of hepatocytes in adult mice. In addition to constitutive transgene expression, this system can be used for more advanced applications, such as shRNA-mediated gene knock-down, implication of the CRISPR/Cas9 system to induce gene mutations, or inducible systems. Here, the combination of constitutive CreER expression together with inducible expression of a transgene or miR-shRNA of choice is presented as an example of this technique. We cover the multi-step procedure starting from the preparation of sleeping beauty-transposon constructs, to the injection and treatment of mice, and the preparation of liver tissue for analysis by immunostaining. The system presented is a reliable and efficient approach to achieve complex genetic manipulations in hepatocytes. It is specifically useful in combination with Cre/loxP-based mouse strains and can be applied to a variety of models in the research of liver disease. |
| Databáze: | OpenAIRE |
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