Virgin collection and haplodiploid crossing methods in Nasonia (parasitoid wasp)

This protocol covers most procedures used by the Nasonia (wasp) geneticist, including virgin collection. Nasonia are easily sexed as pupae, so collecting virgins can easily be performed within a window of several days. The female-specific ovipositor is visible at even the earliest pupal stages as a longitudinal ridge on the ventral posterior abdomen. To prolong generations, yellow pupae can be stored at 4 degrees C-8 degrees C for up to 2 mo and adults for 2-3 wk. In the example presented here, haplodiploid crossing methods are used to make a new Nasonia vitripennis mutant homozygous, using an aunt/nephew mating approach. The protocol can be extended to inbreeding strains. Inbreeding depression generally is not a problem in Nasonia unless the mutation itself is female-sterile. Mother/son mating and sib mating are also reasonable approaches to inbreeding.