Inducing Aiptasia Gamete Release

To induce Aiptasia gamete release following a simulated full moon cue, individuals with oral disk diameter ~0.7 cm or greater were combined into couples ([H2xCC7] or [F003xCC7]), transferred to ASW in small food-grade translucent polycarbonate tanks (GN 1/9–65 cm height, #92 CW; Cambro, Huntington Beach, CA, USA) and kept in standard cell culture incubators (Heracell 150i; Thermo Scientific, Waltham, USA) at 27 °C–30 °C (see temperatures indicated in Results). Tanks were fed Artemia nauplii five times per week and kept on a diurnal 12L:12D cycle at intensities of 20–30 μmol m⁻² s⁻¹ provided by white LED lights, which were comprised of two types of LEDs—those with wavelengths of 425 nm–705 nm (color temperature 15,000 Kelvin) and those with wavelength 460 nm – at a ratio of 5:1 respectively (SolarStinger SunStrip “Marine”; Econlux, Cologne, Germany). A 29-day lunar cycle was simulated by subjecting the tanks to constant blue light during the 12 h “dark” period in the first five nights of the cycle (Day 1–5). Blue LED lights were comprised of LEDs with wavelengths 400 nm, 420 nm, 440 nm, and 460 nm at a ratio of 1:1:1:1 (SolarStinger SunStrip “Deep Blue”; Econlux, Cologne, Germany) and were 10–16 μmol m⁻² s⁻¹ in intensity. The presence of gametes or planula larvae was checked by examining the tanks with a Leica S8APO stereoscope.