GENETIC-VARIATION AMONG CULTIVARS OF RED-CLOVER (TRIFOLIUM-PRATENSE L) DETECTED BY RAPD MARKERS AMPLIFIED FROM BULK GENOMIC DNA

The use of random amplified polymorphic DNA (RAPD) markers obtained fr om bulked samples was investigated for cultivar identification in red clover. Pooled samples were examined in order to minimize variation wi thin cultivars. To determine the appropriate number of individuals to include in the bulked samples representing each cultivar, DNA samples from two, three, four, five, ten and twenty individuals were pooled. T wenty was found to be an appropriate number of red clover individuals per bulk in order to amplify only the DNA sequences shared among most individuals in each cultivar. Fourteen 10-mer primers were used to amp lify genomic DNA from combined leaf samples of 15 red clover cultivars from European, Japanese and North American origins. A total of 79 amp lified products, of which 55 were polymorphic, was obtained. Cultivar- specific bands were observed with 13 primers. The amplification patter ns obtained from two primers could distinguish all 15 red clover culti vars. Rogers' genetic distances for all 105 pairwise comparisons were calculated to evaluate relationships among these cultivars. Cluster an alysis based on these genetic distances separated these 15 cultivars i nto three groups, with two of the groups consisting of a single Japane se cultivar each, while the third group included cultivars from Europe an, North American, and Japanese origins.