Primers were developed for 118 microsatellites isolated from grape (Vitis vinifera) genomic libraries enriched for (AC)n repeats. Only one microsatellite sequence matched other grape SSR-sequences in the GeneBank database. Genotyping was carried out in the parental lines and four offspring of two pseudo-test-cross populations, 'Cabernet Sauvignon' x 'Seyval' and 'Chardonnay' x 'Bianca', and a further six other grape genotypes (V. vinifera 'Sultanina', 'Merlot', 'Syrah', 'Müller-Thurgau', Vitis 'Regent' and V. riparia 'Gloire de Montpellier'). A total of 108 microsatellites showed easily scorable alleles and 100 of them segregated according to a configuration suitable for mapping in either cross. A further 8 SSRs, although unsuitable for mapping in those crosses, showed polymorphism in the other genotypes tested. This set of markers was used, along with 75 microsatellites of other repeat-types, to fingerprint 46 offspring of the cross 'Chardonnay' x 'Bianca'. For each full-sib, individual heterozygosity and distance in repeat units between pairs of alleles at each locus (mean d 2) were calculated as a tool for predicting 'highly outbred' recombinant individuals. Six microsatellites with segregation ratios significantly distorted towards the lack of homozygous sibs were identified and mapped to linkage groups LG 3 and LG 5. Estimation of heterozygosity at genome-wide level and genotyping at loci for which homozygous sibs are discriminated against are discussed for marker-assisted background selection in outcrossing grapevines. © 2005 Springer.