All too often, studies of sexual selection focus exclusively on the responses in one sex, on single traits, typically those that are exaggerated and strongly sexually dimorphic. They ignore a range of less obvious traits and behavior, in both sexes, involved in the interactions leading to mate choice. To remedy this imbalance, we analyze a textbook example of sexual selection in the stalk-eyed fly (Diasemopsis meigenii). We studied several traits in a novel, insightful, and efficient experimental design, examining 2,400 male–female pairs in a “round-robin” array, where each female was tested against multiple males and vice versa. In D. meigenii, females exhibit strong mate preference for males with highly exaggerated eyespan, and so we deliberately constrained variation in male eyespan to reveal the importance of other traits. Males performing more precopulatory behavior were more likely to attempt to mate with females and be accepted by them. However, behavior was not a necessary part of courtship, as it was absent from over almost half the interactions. Males with larger reproductive organs (testes and accessory glands) did not make more mating attempts, but there was a strong tendency for females to accept mating attempts from such males. How females detect differences in male reproductive organ size remains unclear. In addition, females with larger eyespan, an indicator of size and fecundity, attracted more mating attempts from males, but this trait did not alter female acceptance. Genetic variation among males had a strong influence on male mating attempts and female acceptance, both via the traits we studied and other unmeasured attributes. These findings demonstrate the importance of assaying multiple traits in males and females, rather than focusing solely on prominent and exaggerated sexually dimorphic traits. The approach allows a more complete understanding of the complex mating decisions made by both males and females.

2400 male-female mating pairs outcomesMales&Females.xlsx This file contains all information of each male-female pair, along with PCA for reproductive organs and behavior. The columns are labelled: Line – inbred line (12) in.out – inbred/outbred status (outbred all belong to one line) Male.ES – male eyespan Male.Thorax Testes AG – Accessory Glands Female.ID Female.ES – female eyespan Accept.Reject – given that a mating attempt occurred Male.ID Attempt – was there a mating attempt, yes or no Follow – a behaviour Bob – a behavior Grapple – a behavior Time.to.mate – how many seconds untila mating attempt Date – block structure Time.in.pair – same as Time.to.mate except that those where there was no mating attempt are labelled 900 seconds PC.Reporgan – PCA Testes and Accessory Glands PC1Beh.min – PCA of three behaviours per minute PC2Beh.min - PCA of three behaviours per minuteMales&Females.xlsxMaleSummaryThis file is an abstraction from the larger file Males&Females.xlsx, summarizing for each male. Behaviour is not recorded here as it relates to the particular pair.