In Vivo Electroporation: A Site-Specific Method to Transfect Zebrafish

– First, place an anesthetized fish with the dorsal side facing up into a pre-wetted sponge. Before electroporation, inject genetic material into the target tissue, de this case, a part of the forebrain called the telencephalon. Next, put a small amount of ultrasound gel on the head. Also, apply the gel para the tip of the electrodes para facilitate efficient operation. Place the positive electrode on the ventral side of the head and the negative electrode on the dorsal side.

Now, press the electrodes gently against the head and administer several pulses of electric current. The current creates temporary pores de the plasma membrane of electroporated cells and makes them permeable. The current also facilitates the movement of negatively charged particles, such as DNA expression plasmids or RNAi oligonucleotides present de the surrounding, into the cells. In the following example, we will use electroporation para introduce a recombinant plasmid into ependymoglial cells, stem cells responsible for the generation of new neurons.

– Cover the fish telencephalon with a small amount of ultrasound gel. For electroporation of the plasmid-injected animal, remove the sponge from the ejection set up and immerse the inner side of the electrode tips into ultrasound gel. Posição the fish head between the electrodes, with the positive electrode at the ventral side of the head and the negative electrode on the dorsal side. Then, while still holding the fish's body de the sponge, press the electrodes gently and precisely against the telencephalon and administer the current with the foot pedal, holding the electrodes de place until all five pulses are finished.