Although the process of double fertilization is a defining feature of angiosperms, it has only been carefully documented in relatively few, mostly derived, angiosperm species. Recent phylogenetic analyses have placed Nymphaeaceae as either the first or second diverging lineage from the base of the angiosperm tree. We studied the fertilization biology of Nuphar polysepalum (Nymphaeaceae). Here we confirm for the first time, using modern tools, the presence of double fertilization in an angiosperm basal to the monocot and eudicot clades. Another important and presumed synapomorphy of angiosperms, virtually unstudied in early angiosperms, is the highly reduced, monosporic eight-nucleate female gametophyte (embryo sac). The second fertilization in such an embryo sac is expected to produce a triploid primary endosperm nucleus, via the fusion of two female polar nuclei and one male sperm nucleus. Interestingly, the mature Nuphar embryo sac consists of only four nuclei: two synergids, an egg, and one polar, or central cell, nucleus. We confirmed the haploid nature of the central cell nucleus using DNA quantitation of individual nuclei. The central cell nucleus has the same DNA content as known haploid nuclei, such as the egg. We further show that, in cross-pollinated flowers, the products of double fertilization, the zygote nucleus and the primary endosperm nucleus, have equal initial DNA quantities, instead of the 2:3 ratio. Thus, contrary to expectation, the endosperm in one of the earliest angiosperm lineages is biparental and diploid, not triploid. The lack of information regarding basic reproductive biology in basal angiosperms represents a huge gap in our knowledge that may hold further surprises.

Key words: basal angiosperms, character evolution, double fertilization, embryology, female gametophyte, Nymphaceae