Twenty-one porcine microsatellite markers were developed by screening DNA libraries and by a computer search of databases. The microsatellites were typed in a large three-generation family established by a cross between the European wild pig and a Swedish Yorkshire breed. Linkage analysis benefited from the fact that due to the divergence between the parental populations, the degree of microsatellite polymorphism was significantly higher in the F1 animals than in either of the parental populations. Parallel typing of a set of 35 restriction fragment length polymorphism, protein, and blood group markers rendered it possible to assign as many as 20 of the microsatellites to the porcine linkage map. Fourteen microsatellites were localized to a chromosome segment, whereas six constituted parts of unassigned linkage groups. Analysis of four microsatellites within genes allowed the assignment of the endoplasmic reticulum Ca2+ transport ATPase locus to chromosome 14, the assignment of the interferon-gamma and the diacylglycerol kinase loci to a new linkage group (XI), and the localization of the tumor necrosis factor beta locus close to the major histocompatibility complex (SLA) on chromosome 7 to be confirmed. Fluorescence in situ hybridization mapping of two microsatellite-containing cosmids assigned two linkage groups to chromosomes 9 and 12, respectively. In total, 27 new markers were added to the porcine linkage map, thereby almost doubling the number of markers on the map. Linkage groups are now present on 10 of 18 of the pig autosomes.(ABSTRACT TRUNCATED AT 250 WORDS)