Abstract
Accessions with no germinating seeds are a common problem in plant gene banks and research institutions. Our goal was to elaborate and apply an in vitro method of germination and multiplication of old aged seeds of red and alsike clover and alfalfa. Eighteen clover and five alfalfa accessions were used for germination in vitro. Most of the accessions had produced seeds more than 20 years ago and the seeds did not germinate in soil. Seed pre-treatment with different concentrations of potassium permanganate, as well as addition of phytohormones, AgNO3 and activated carbon to germinating media were tested. Plantlets for all germinated accessions were obtained, even in the case when seeds were 44-year-old (alfalfa). Germination rate in vitro not always correlated with seed age and ranged from 2 to 72%. Pre-treatment with potassium permanganate was effective both for seeds sterilisation and germination stimulation. Most germinated seeds formed phenotypically normal seedlings with all organs. In vitro multiplication of obtained clover plants was performed. Best results were achieved by micropropagation of stem segments approximately 2 mm in length. Number of finally obtained plants depended not only on seed germination ability, but also on micropropagation ability in relation to genotype, and on acclimatization success in soil after in vitro cultivation