A study of genetic diversity in Trifolium hybridum varieties using morphological characters and ISSR markers

The nutritive value of alsike clover (Trifolium hybridum L.) is slightly lower than that of red and white clovers; however, it exhibits some advantages – persists longer in a sward, performs well on acid and wet soils, suits for forage and silage production, can be used as green manure, and is a melliferous plant. With declining liming volumes, the soils are rapidly acidifying, which may result in an increasing alsike clover production area. To reveal plant diversity, the methods currently employed encompass not only research into agro-biological traits but also biochemical-molecular assays. The inter-simple sequence repeats (ISSR) technique is one of the DNA assay methods widely used in genetic diversity studies, genetic evolution and systematic specification, and gene mapping. The current study was aimed to assess individual plants of T. hybridum varieties (‘Daubiai’, ‘Lomiai’, ‘Poliai’, ‘Skriveru’) for morphological traits, quality parameters and genetic diversity and to establish the correlations between the ISSR markers and characteristics of agro-biological traits. Morphological diversity was estimated according to 10 traits. The coefficient of variation varied within a wide range (CV 0.6–71.9%). The parameters of feeding value were found to be most stable (CV 0.6–16.6%), while seed yield was the most variable indicator (CV 38.7–71.9%). According to the values of agro-biological traits, the individual plants conditionally split into 4 sub-clusters, encompassing different varieties. Five primers were used to reveal the DNA diversity of alsike clover. A total of 37 fragments were amplified, of which 62.2% were polymorphic. Genetically most distant were individuals of ‘Lomiai’ and ‘Daubiai’ cultivars. The dendrograms produced according to the values of agro-biological and genetic distances were not identical. The ISSR markers were found to correlate with the following alsike clover traits: seed yield, stem number and height, crude protein and water soluble carbohydrate contents.