One hypothesis derived from evolutionary perspectives is that men are more upset than women by sexual infidelity and women are more upset than men by emotional infidelity. The proposed explanation is that men, in contrast to women, face the risk of unwittingly investing in genetically unrelated offspring. Most studies, however, have relied on small college or community samples of heterosexual participants. We examined upset over sexual versus emotional jealousy among 63,894 gay, lesbian, bisexual, and heterosexual participants. Participants imagined which would upset them more: their partners having sex with someone else (but not falling in love with them) or their partners falling in love with someone else (but not having sex with them). Consistent with this evolutionary perspective, heterosexual men were more likely than heterosexual women to be upset by sexual infidelity (54 vs. 35 %) and less likely than heterosexual women to be upset by emotional infidelity (46 vs. 65 %). This gender difference emerged across age groups, income levels, history of being cheated on, history of being unfaithful, relationship type, and length. The gender difference, however, was limited to heterosexual participants. Bisexual men and women did not differ significantly from each other in upset over sexual infidelity (30 vs. 27 %), regardless of whether they were currently dating a man (35 vs. 29 %) or woman (28 vs. 20 %). Gay men and lesbian women also did not differ (32 vs. 34 %). The findings present strong evidence that a gender difference exists in a broad sample of U.S. adults, but only among heterosexuals.