Plant Regeneration from Leaf-derived Callus Cultures of Primrose (Primula vulgaris)

Abstract

Efficient micropropagation of Primula species is important both for fundamental scientific studies and commercial applications. Primula vulgaris (Huds), along with other Primulaceae species, exhibits floral heteromorphy with two distinct forms of hermaphroditic flower. Studies to identify genes that control heteromorphic flower development require propagation of floral mutants, and efficient regeneration is a key requirement for plant transformation. Several species, including P. vulgaris cultivars and P. ×polyantha hybrids, are important horticultural crops in Europe, United States, and Japan and semidouble/double Primula varieties offer a high-end product. Vegetative propagation of sterile double forms, and as a means to increase numbers of inbred parent plants for F1 seed production is, however, slow. Micropropagation offers the most efficient way of increasing these varieties quickly and efficiently. To date, most Primula micropropagation protocols require explant material derived from in vitro grown seedlings or use floral parts as donor material with seasonal limitations. Therefore, an effective and efficient protocol was developed for in vitro regeneration of P. vulgaris via indirect organogenesis from adult leaf–derived explants. Exposure of leaf explants of P. vulgaris to media containing synthetic cytokinin, thidiazuron (TDZ), and auxin [1-naphthylacetic acid (NAA) or 2,4-dichlorophenoxyacetic acid (2,4-D)] resulted in undifferentiated cell proliferation and followed by differentiated growth as shoot organogenesis. Silver nitrate improved in vitro callus growth and increased shoot regeneration further, with up to 72% of explants producing shoots. Regenerated plants developed normally and produced normal fertile flowers within 7 months. The system was also successfully applied for the micropropagation of sterile double-flowered P. vulgaris ‘Sue Jervis’. The protocol reported here enables propagation of P. vulgaris without seasonal limitation or destruction of valuable parent donor material. The protocol, with further development, has the potential to underpin development of a transformation system for Primula, which would be of value in studies on flower development and disease resistance in laboratory grown plants.