Hi all, a common issue I face when making new strains such as double mutants is that my final plate containing the desired strain also shows a good number of male population. I use him-5 males to outcross at the beginning. Is having males on plate a problem, esp if I do assays such as percentage L1s surviving to adulthood, or washing worms off the plate to do rt-qpcr with wild-type N2 worms - that contains much fewer males - as the control? Thank you for your insight into this matter.
Yes, as you suspected, him-5 is something you really should get rid of in your final strain since it is going to affect brood sizes and continue generating pesky males, XXX hermaphrodites and embryonic lethals. It is a nuisance since you have to clone about 12 L4 hermaphrodites, wait for the F2 to come up and then discard any plates that have obvious males present (if in a rush, you could do single worm PCR and check for e1490 or whatever allele you are using). Him strains are convenient for lots of things, but in most cases you would probably be better off using N2 males for constructions (or using heat shock or him RNAi etc).