Lethal phase determination

Lethal phase determination

 

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1. Pick a bunch of heterozygote L4 hermaphrodites to a new plate.

2. The next day, there will be eggs on the plate; pick exactly 20 of these to each of a few plates. Make sure to use first day eggs like this: wild-type animals eventually run out of sperm and lay unfertilized eggs that don't develop, and you want to avoid confusing these with dead fertilized eggs. Line up the sets of 20 eggs in a row just beyond the edge of the seeded portion of the plate.

3. The next day and succeeding days, examine the plates for unhatched eggs, dead worms, and live worms.

4. If the worms hatch, but may be dying as young larvae, it can be difficult to distinguish between worms being "gone" because they died and the dead larvae are hard to find, or because live worms just crawled up the side of the dish and disappeared. To help this problem, you can keep the worms from crawling off the bacterial lawn by using a P-20 to ring the lawn with a line of 10 mg/ml palmitic acid in EtOH, which precipitates on the plate and forms a barrier the worms won't cross.