Plant regeneration via embryo-like structures: histological observations and genetic stability in regenerants of Lilium spp.

Abstract

SUMMARYTransverse thin-cell layers (tTCLs) excised from bulblets of Lilium longiflorum X Oriental hybrid ‘Triumphator’ were cultured on 1.0X Murashige and Skoog (MS) medium containing 0.1 mg l−1 α-naphthaleneacetic acid (NAA) and 0.1 mg l−1 kinetin (KT) in the dark at 23° ± 2°C. White-yellow, friable, embryo-like structures formed after 6 weeks in culture. The embryo-like structures proliferated well when cultured on 1.0X MS medium containing 1.0 mg l−1 NAA and 0.2 mg l−1 thidiazuron (TDZ), and converted into whole plantlets on 0.5X MS medium without any added plant growth regulator. Histological studies identified the origin of the first cell divisions from single sub-epidermal cells, from which somatic embryos developed. Using this procedure, embryo-like structures (ELS) were obtained at frequencies of 85.5%, 76.8%, 68.4%, 28.5% and 25.6% for L. longiflorum, L. longiflorum X Oriental ‘Triumphator’, Lilium Oriental hybrid ‘Siberia’, Lilium Asiatic hybrid ‘Elite’, and L. davidii var. unicolor, respectively. Embryo-like structures could be proliferated and developed into plantlets, with five-to-ten plantlets per embryo-like structure in all five Lilium species and hybrids tested. No polymorphic bands were detected using inter-simple sequence repeat (ISSR) markers and no change in ploidy level was found by flow cytometry (FCM) among the regenerants of all five Lilium species and hybrids. These five Lilium species and hybrids represented a diverse genetic resource of the genus Lilium. This protocol may be widely applicable for somatic embryogenesis and has potential applications in micropropagation and genetic transformation in Lilium spp.