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• W2080392322 abstract "This video and article contribution gives a comprehensive description of microinjection and electroporation of mouse testis in vivo. This particular transfection technique for testicular mouse cells allows the study of unique processes in spermatogenesis. The following protocol focuses on transfection of testicular mouse cells with plasmid constructs. Specifically, we used the reporter vector pEGFP-C1, which expresses enhanced green fluorescent protein (eGFP) and also the pDsRed2-N1 vector expressing red fluorescent protein (DsRed2). Both encoded reporter genes were under the control of the human cytomegalovirus immediate-early promoter (CMV). For performing gene transfer into mouse testes, the reporter plasmid constructs are injected into testes of living mice. To that end, the testis of an anaesthetized animal is exposed and the site of microinjection is prepared. Our preferred place of injection is the efferent duct, with the ultimately connected rete testis as the anatomical transport route of the spermatozoa between the testis and the epididymis. In this way, the filling of the seminiferous tubules after microinjection is excellently managed and controlled due to the use of stained DNA solutions. After observing a sufficient filling of the testis by its colored tubule structure, the organ is electroporated. This enables the transfer of the DNA solution into the testicular cells. Following 3 days of incubation, the testis is removed and investigated under the microscope for green or red fluorescence, illustrating transfection success. Generally, this protocol can be employed for delivering DNA- or RNA- constructs into living mouse testis in order to (over)express or knock down genes, facilitating in vivo gene function analysis. Furthermore, it is suitable for studying reporter constructs or putative gene regulatory elements. Thus, the main advantages of the electroporation technique are fast performance in combination with low effort as well as the moderate technical equipment and skills required compared to alternative techniques." @default.
• W2080392322 created "2016-06-24" @default.
• W2080392322 creator A5040985484 @default.
• W2080392322 creator A5069867918 @default.
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• W2080392322 date "2014-08-23" @default.
• W2080392322 modified "2023-10-03" @default.
• W2080392322 title "<em>In Vivo</em> Microinjection and Electroporation of Mouse Testis" @default.
• W2080392322 cites W1536918519 @default.
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• W2080392322 doi "https://doi.org/10.3791/51802" @default.
• W2080392322 hasPubMedCentralId "https://www.ncbi.nlm.nih.gov/pmc/articles/4758768" @default.
• W2080392322 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/25177859" @default.
• W2080392322 hasPublicationYear "2014" @default.
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