We conducted a factorial field experiment in order to investigate the role of the soil seed bank in gap recolonization in a calcareous grassland, using 240 experimental gaps (10 x 10 cm). Experimental manipulations included the elimination of the seed bank (by sterilizing the soil), the elimination of short-distance seed rain by removing all flowerheads in 2 x 2 m plots, and the prevention of lateral clonal spread by surrounding gap soil with 41-μm nylon mesh. The influence of competition on regeneration was also studied, by planting an adult individual of Filipendula vulgaris or Festuca rubra in the center of some gaps. There were 130 species of vascular plants in the established vegetation. Twenty-five species also occurred as seedlings; 51 species were recorded in the seed bank, of which only two were not present in the growing vegetation. There were on average 2362 seeds/m2 in a 10 cm deep soil layer. Species relative abundances in the established vegetation and in the seed bank were not significantly correlated. Eighty-four vascular plant species emerged from seeds in the experimental gaps. The seed bank contributed on average 5.4 seedlings per gap, short-distance seed rain (<0.5 m) 4.1 seedlings, and longer distance (>0.5 m) seed rain 2.7 seedlings. Thirty-one vascular plant species arrived in experimental gaps by lateral clonal spread. Of these, only one was not recorded as a seedling in the experimental gaps. There were on average 2.7 clonally spread shoots per gap. Species with smaller seeds were present in significantly greater numbers in the seed bank than in the vegetation and were also more common colonizers of experimental gaps. There was no evidence of competition or facilitation between seedlings, or between seedlings and adults. It was concluded that the soil seed bank has an important functional role in a perennial grassland community as a means for population maintenance for many species. Thirty-six percent of the regeneration in small gaps was due to the seed bank, and one may expect that this proportion will increase in larger gaps.