Poplars (Populus sp.) are among the strongest isoprene (Iso)-emitting plants. Ten poplar genotypes belonging to four different species were grown under the same environmental conditions in a common garden experiment, to study the influence of the genetic variability on Iso emission and on the relationship between Iso and photosynthesis. Photosynthesis ranged from 13 to 20 µmol CO2 m−2 s−1, whereas Iso emission ranged from 18.2 to 45.2 nmol m−2 s−1. There was no clear association between Iso emission and photosynthesis. In most genotypes, photosynthetic capacity developed earlier than Iso emission capacity. The emission of Iso was inversely correlated with the intercellular CO2 concentration (Ci) and positively correlated with instantaneous water use efficiency. It is speculated that, by regulating Ci, stomatal opening also indirectly controls Iso emission in poplars. A positive linear correlation between the fraction of recently assimilated carbon emitted as Iso and Iso emission rate was found. The slope of this relationship indicated that each nanomole of Iso emitted requires a fixed fraction of photosynthetic carbon regardless of the intra- and interspecific variability in the Populus genus, and of leaf ontogeny. A comparison with data from recent studies showed that the slope of this relationship increases in drought-stressed leaves. However, this might be explained by an increasing contribution of carbon sources for Iso biosynthesis from stored photosynthates. If this is true, then the amount of carbon directly shunted from photosynthesis into Iso is constant in all poplars and is not influenced by abiotic stresses.