Lethalphasedetermination

1.PickabunchofheterozygoteL4hermaphroditestoanewplate.

2.Thenextday,therewillbeeggsontheplate;pickexactly20ofthesetoeachofafewplates.Makesuretousefirstdayeggslikethis:wild-typeanimalseventuallyrunoutofspermandlayunfertilizedeggsthatdon"tdevelop,andyouwanttoavoidconfusingthesewithdeadfertilizedeggs.Lineupthesetsof20eggsinarowjustbeyondtheedgeoftheseededportionoftheplate.

3.Thenextdayandsucceedingdays,examinetheplatesforunhatchedeggs,deadworms,andliveworms.

4.Ifthewormshatch,butmaybedyingasyounglarvae,itcanbedifficulttodistinguishbetweenwormsbeing"gone"becausetheydiedandthedeadlarvaearehardtofind,orbecauselivewormsjustcrawledupthesideofthedishanddisappeared.Tohelpthisproblem,youcankeepthewormsfromcrawlingoffthebacteriallawnbyusingaP-20toringthelawnwithalineof10mg/mlpalmiticacidinEtOH,whichprecipitatesontheplateandformsabarrierthewormswon"tcross.

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