Testes measures of sterile and fertile males
Males from each strain as well as sterile hybrid males were collected and kept in vials containing CYAM medium with no more than 20 males per vial to avoid crowding. The males were aged to either 5 or 10 days old before being frozen to preserve them. Frozen males were allowed to briefly thaw over the course of a few minutes and dissected in a drop of 1× PBS. Forceps were used to grip the thorax while the other pinched the male’s abdomen just above the aedeagus. Then, by simply pulling on the gripped abdomen the entire male’s reproductive tract was pulled out into a drop of 1×PBS. The testes were isolated using dissecting pins and moved onto a fresh drop of 1×PBS. Testes images were captured using an inverted Olympus CKX41 microscope and the image processing and analysis software Image J (https://imagej.nih.gov/ij/) was used to measure an area towards the basal end of the testes where mature sperm is found and which we refer to as the seminal vesicle. The area can be approximately identified by an apical pinch followed by a basal pinch or torsion. One testis from each male was measured.