Micropropagation of common ash clones resistant to fungus Hymenoscyphus fraxineus

Abstract

A regeneration system was studied in Fraxinus excelsior from the juvenile and mature plant material. Different explants were used for the in vitro establishment. Plant material was cultured on full-strength Woody Plant Medium (WPM) supplemented with 0.5 mg L-1 N6-(3-hydroxybenzyl)adenine (meta-topolin) when mature explants (stems) were used or without plant growth regulators when juvenile explants (seeds and embryos) were considered. The best regeneration rate was observed for zygotic embryos where 98.5% embryos developed into seedlings. Extraction of embryos from seeds significantly decreased contamination rate and increased development rate compared to complete seeds. When stem explants were used, differences in contamination and regeneration among various types of explants were recorded. Apical segments with one tip and excised apical tips with leaf primordia appeared 50% free from fungal and bacterial contaminations. All intact apical segments were lost by 100% contamination. Proliferation was higher in apical segments with one tip compared to excised apical tips. The presence of antibiotic solution in medium had mostly no significant effect on reduction of explant contamination and on shoot proliferation compared to the treatment without antibiotics. The only exception was the initial culture established from the Novosedlo locality where antibiotics had negative effect.