Segregation distortion (SD) is the deviation of genetic segregation ratios from their expected Mendelian fraction and is a common phenomenon found in most genetic mapping studies. In this study two segregating Lolium perenne populations were used to construct two genetic maps: an 'F(2) biomass' consisting of 360 genotypes and an 'F(1) late flowering' sibling based population consisting of 182 genotypes. Additionally two parental maps were generated for the 'F(1) late flowering' population. SD was detected and p-values for SD were calculated for each marker locus. The 'F(1) late flowering' map had only half of the extent of SD (32%) compared to the map based on the 'F(2) biomass' population (63%). Molecular marker data have been supplemented with genomic in situ hybridization (GISH) data to show non major non-recombined segments of Fescue chromosomes within the parental inbred ryegrass lines with a Festuca x Lolium pedigree. We conclude that SD in our study is more likely caused by genetic effects rather than by population structure and marker types. Two new L. perenne mapping populations including their genetic maps are introduced; one of them is the largest reported Lolium mapping population consisting of 360 individuals.