In Denmark, two seemingly distinct size-classes, 3^rd and 4^th instar juveniles and reproductive adults, of Araneus diadematus are found during every breeding season in autumn, indicating a non-overlapping biennial life-cycle. We tested the hypothesis that alternate generations might experience a degree of reproductive isolation, using the distribution of nuclear (allozyme) and maternal (mtDNA) genetic markers. Individuals of a locality behaved as belonging to a random mating population, irrespective of size. No differences were found between any size-class pairs, within and between 2 yr, or among geographically distant samples. Processes that may lead to this result are discussed: the biennial development may be incomplete; or there may be migrational influx of genes from southern annual populations. There is no evidence for sexual differences in life-cycle length.