Abstract
The genus Helleborus comprises 22 species, which are allocated to six Helleborus sections. Helleborus species show differences with regard to leaf and flower morphology, scent, and sensitivity to different pests and diseases. Breeding programs aiming at these traits require the inclusion of a broader spectrum of Helleborus species in addition to the most popular species H. niger (Christmas Rose) and H. A hybridus (Lenten Rose), thus interspecific hybridisations need to be performed. Pollen viability was estimated for nine species by staining with thiazolyl blue (MTT) and varied between 67 and 99% for fresh pollen and between 74 and 99% for pollen dried for 24 h. Crossing barriers had previously been localized in Helleborus species as predominantly post-zygotic. Therefore, embryo rescue techniques via ovule culture were established to overcome these barriers. Ovules were isolated from the maternal plants five to seven weeks after pollination and then cultured in vitro. Overall, from 661 crosses involving 13 Helleborus species, 40,885 ovules were obtained. In a combination with parents of low genetic distances between the parental species, up to 26.08% of these ovules produced hybrid shoots, whereas in intersectional crosses this was the case in only 0.17 to 0.85% of the ovules. In total, 217 hybrid offspring were verified by flow cytometry and molecular marker analysis (random amplified polymorphic DNAs), whereof 16 were derived from parental species belonging to different Helleborus sections. Hybrid plantlets of some cross combinations were transferred to the greenhouse and expressed intermediate leaf and flower morphology when compared to the parental species.
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