Ploidy level stability of adventitious shoots of sour cherry ‘Čačanski Rubin’ and Gisela 5 cherry rootstock

Abstract

The capacity of regeneration of adventitious shoots from leaf explants was studied in sour cherry ‘Cacanski Rubin’ (Prunus cerasus L.) and cherry rootstock Gisela 5 (P. cerasus × P. canescens). Regeneration assay included thirty different combinations of plant growth regulators. 6-benzyladenine (BA) and thidiazuron (TDZ) were applied either individually or each combined with different concentrations of indole-3-butyric acid, α-naphthaleneacetic acid (NAA) and 2,4-dichlorophenoxyacetic acid (2,4-D). ‘Cacanski Rubin’ showed higher regeneration capacity in comparison with Gisela 5 regarding the total number of treatments inducing regeneration as well as the highest frequency of regeneration achieved. In both genotypes, 8.9 μM BA was more effective than both 4.5 and 9.0 μM TDZ in inducing adventitious regeneration, but only when combined with auxins. The highest frequency of regeneration (20.8 %) in ‘Cacanski Rubin’ was achieved on medium supplemented with 8.9 μM BA combined with 5.4 μM NAA, while in Gisela 5 the highest value (8.3 %) was obtained when BA was combined with 4.5 μM 2,4-D. Flow cytometry combined with 4′-6-diamidino-2-phenylindole staining was employed to estimate DNA ploidy levels and relative nuclear DNA content in adventitious regeneration-derived shoots, in vitro shoots of axillary origin and in vivo control plants from open field. No significant differences in nuclear DNA content were detected among plants of different origin. Chromosome counting in root tip meristems also showed normal tetraploid chromosome number (2n = 4x = 32) in ‘Cacanski Rubin’ shoots and normal triploid chromosome number (2n = 3x = 24) in Gisela 5 shoots regenerated in vitro. The results obtained suggest that no major genetic instability occurred during adventitious regeneration under the described experimental conditions.