Perennial ryegrass (Lolium perenne L.) is one of the most important forage grasses, providing high yields with excellent forage quality. The main limiting factor for increasing the cultivation area of perennial ryegrass in the Nordic-Baltic region is insufficient winter hardiness due to unstable climatic conditions as well as insufficient persistence and drought resistance. Currently, the genetic diversity of perennial ryegrass cultivars is relatively limited; therefore, developing new, highly adaptable germplasm is of high importance in the context of changing climatic conditions. In the framework of the Nordic-Baltic Public-Private Partnership (PPP) project in pre-breeding of perennial ryegrass, 250 tetraploid plants (hereinafter genotypes), created by chromosome doubling using colchicine at the Institute of Agriculture, Lithuanian Research Centre for Agriculture and Forestry, were evaluated in open field conditions at the Research Institute of Agronomy of Latvia University of Life Sciences and Technologies. Detailed phenological scoring of all genotypes was performed over a three-year (2016-2018) period. For data analysis, the plants were grouped according to heading time and growth habit. Significant differences among the groups were found in winterhardiness, regrowth rate, development rate of generative shoots, susceptibility to rust, etc. The seed from plants that survived well and showed some promising properties were harvested in the 2 nd ley year-a total of 199 genotypes or 358 individuals (80% and 48% of all, respectively). Results of genotyping of randomly selected genotypes with the highest and lowest winter hardiness showed that the clones were genetically differentiated from the cultivars developed in Baltic countries-unique alleles were found in the tetraploid clones that were not present in the analysed cultivars. This suggests that these developed tetraploid clones or genotypes could provide valuable breeding material to improve the suitability of perennial grass cultivars to local environmental conditions in the future.