The ‘living fossil’ Ginkgo biloba L. is the only extant representative of Ginkgophyta, which is an ancient group of gymnosperms that constituted an important component of the Earth’s forests in the Mesozoic and early to mid-Cenozoic. The Ginkgo and its sister phylogenetic relatives, the cycads (Cycadophyta), are the last major lineages of green plants in which polyploidy (whole genome duplication) remains unknown. Surprisingly, however, Ginkgo has the potential to form spontaneous polyploid offspring. We found such a vital polyploid sapling of Ginkgo during a routine screening for genome size variation in plants used for cultivation experiments. This polyploid sapling (sex yet unknown) originated from the seeds collected from three female trees grown in the Botanical Garden of the Faculty of Science, Masaryk University in Brno (Czech Republic). Its genome size (2C = 37.4 0.2 Gbp) is approximately double that of the diploid Ginkgo biloba (2C = 18.4 0.1 Gbp, mean of all three possible mother trees), indicating that it is tetraploid. Compared with its diploid parental plants and same-age siblings, the leaves of the tetraploid had finely laciniate distal margins and enlarged stomata: 60 ± 6 µm in the tetraploid sapling vs. 39 ± 5 µm in its same-age diploid siblings or 34 ± 6 µm in putatively parental trees.