Chick Embryo Electroporation

Incubate egg on side for 50 h to electroporate stage 12/13 embryos.

Prepare eggs:
1. Punch a hole in the egg on the side facing up, near the small end. Don't rotate the egg.
2. Insert and 18 g needle with 9 ml syringe into the egg and withdraw 3 ml egg white.

Electroporation setup:
3. Pour L15 into a small petri dish and into a Falcon tube. Gently clean electrodes in the L15 dish with a brush. Set Electro-porator machine to 25V, 5 pulses of 50 msec.
4. Select a capillary and open the pulled tip by touching it to side of forceps.
5. Insert the non-pulled end of the capillary into the mouth pipet and fill with DNA/Fast Green.

DNA injection and Electroporation:
6. Cut a small window in egg starting at punch hole. Locate the embryo and expand hole towards location of embryo.
7. Point head towards you and note the stage of the embryo.
8. Insert the capillary with DNA into neural tube at a low angle and inject DNA, filling the neural tube.
9. Position embryo for electroporation with head towards electrodes.
10. Under low magnification, position electrodes around the embryo parallel to neural tube. Lower electrodes into egg.
11. Electroporate. (Bubbles should form on electrodes).
12. Add ~4 drops of L15 to the egg (this could also be done before electroporation).
13. Carefully remove electrodes from egg and create an airtight seal with tape.
14. Clean electrodes with L15 before electroporating next egg.
15. When all eggs are done, place tray back into incubator.

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