Obtaining Eggs from Xenopus Laevis Females

Set up a tank containing 100 mM sodium chloride and house frogs at approximately one frog per liter at 18°C. Inject mature female frogs subcutaneously into the dorsal lymph sack with 5 mL of 200 units/mL pregnant mare serum gonadotropin (PMSG), then return them to the sodium chloride tank.

After 2-3 days, inject each frog with 0.5 mL of 100 units/mL human chorionic gonadotropin (hCG) subcutaneously into the dorsal lymph sack. Place each primed frog individually in 5 liters of 100 mM sodium chloride solution.

After 16-22 hours, remove the frogs to a clean water tank and pour the freshly laid eggs into a beaker. Observe frogs for 1-2 days for signs of infection at the injection site.

Rinse eggs once in 2% cysteine pH 8, then incubate in 100 mL of 2% cysteine solution per frog for approximately 5 minutes at room temperature, gently swirling periodically. Pour off the cysteine solution and discard.

Wash eggs three times in 2.5X modified Ringer solution to remove debris, then pour into a 100 mm glass petri dish. Under a dissecting microscope, remove bad eggs (oocytes without white spot, white/enlarged/unevenly pigmented eggs) using a Pasteur pipette, retaining only mature eggs with dark pigmentation and a visible white maturation spot.

Wash the sorted eggs 2-3 times by carefully pouring egg lysis buffer over them and then pouring the buffer off while minimizing spillage of eggs. Eggs are now ready for lysis and extract preparation.