Biotechnology of the rose : a review of recent progress

High yields of protoplasts were isolated from embryogenic suspension cultures of Rosa persica x xanthina and Rosa wichuraiana using an enzyme mixture comprising 20 g l-1 cellulase ‘Onozuka’ R10, 1 g l-1 Pectolyase Y-23 and 10 g l-1 hemicellulase. Agarose-immobilized protoplasts gave the most consistent growth at a plating density of 5×104 protoplasts ml-1 on the basic medium of Kao & Michayluk (KM8p) containing 2 mg l-1 naphthaleneacetic acid and 1 mg l-1 benzylaminopurine. At 25°C in the dark, 0.004% of R. persica x xanthina protoplasts developed into colonies. Using similar culture conditions, but with a plating density of 9×104 protoplasts ml-1, 0.017% of R. wichuraiana protoplasts developed into colonies. On transfer of R. persica x xanthina colonies to Schenk & Hildebrandt's medium containing 3 mg l-1 2,4-dichlorophenoxyacetic acid, globular and later stage embryos were formed. Approximately 30% of these embryos developed into plantlets on transfer to basal Schenk & Hildebrandt's medium. Further development of the plantlets took place on cellulose plugs (Sorbarods) soaked in Murashige & Skoog's medium containing 0.05 mg l-1 naphthaleneacetic acid, 0.05 mg l-1 indole-3-butyric acid and 0.1 mg l-1 benzylaminopurine. Rose breeding is now open to the full range of in vitro genetic manipulation techniques involving protoplast technology.

A protoplast to plant system in roses

Culture conditions are described for sustained cell division and plant regeneration from protoplasts of rose (Rosa hybrida L. `Sumpath'). Protoplasts were enzymatically isolated from 2-week-old embryogenic cell suspension cultures. Freshly isolated protoplasts were plated as a thin layer onto protoplast culture medium (half-strength 21 Murashige and Skoog's medium containing 60 g l−1 myo-inositol, 4.4 μM BA, and 1.4 μM 2,4-D) at a density of 5×104 protoplasts ml−1. The plating efficiency reached 3.9% after 2 weeks of culture. However, few protoplasts underwent cell division when cultured in protoplast culture medium in which 60 g l−1 myo-inositol was replaced with the same osmolarity of 90 g l−1 mannitol, indicating that myo-inositol is essential for sustained cell division of protoplasts. Colonies were formed after 8 weeks of culture at a frequency of 0.2%. Colonies were then transferred to colony culture medium (0.4% Gelrite-solidified protoplast culture medium) and maintained by subculturing at 4-week intervals to form embryogenic calluses. Upon transfer to half-strength MS basal medium, embryogenic calluses gave rise to numerous somatic embryos. Somatic embryos were transferred to half-strength MS basal medium containing 48 mg l−1 ferric ethylenediamine di-(o-hydroxyphenylacetate), where they subsequently developed into plantlets at a frequency of 30.9%. The plantlets had the same chromosome number of 2n=3x=21 as the source plant. They were successfully transplanted to potting soil and grown to maturity in a greenhouse.

Plant regeneration of rose ( Rosa hybridia ) from embryogenic cell-derived protoplasts

The genus Rosa includes over 100 species which are distributed throughout the temperate and subtropical regions of the Northern Hemisphere (Rehder 1960). Chromosome numbers range from 2n = 2x = 14, to 2n = 8x = 56 (Darlington and Wylie 1955). The DNA content of the rose genome is amongst the lowest recorded in the Angiospermae, the 4C value of R. wichuraiana (2n =14) measuring only 0.45–0.48 pg (Lloyd 1986). The chromosomes are correspondingly small (Fig. 1).

Rosa spp. (Roses): In Vitro Culture, Micropropagation, and the Production of Secondary Products

Roses (Rosa spp.) are one of the most important flower crops in the world and have an economic value in ornamental, pharmaceutical and cosmetic trade. Significant progress has been made in biotechnology of roses due to its many potential and practical applications in commercial production and in breeding of roses. Rapid multiplication and production of disease-free plants in vitro have played a vital role in propagation of commercial rose cultivars. Genetic transformation is emerged as an alternative promising tool in rose breeding since it eliminates the difficulties associated with sexual hybridization such as lengthy breeding cycles, sterility, polyploidy and high level of heterozygosity. Biotechnology also allows chimeral segregation and can overcome some of the sterility problems through embryo rescue. In vitro seed germination protocols are ways to shorten breeding cycles and could be used to germinate the seeds that are not possible to germinate by other means. In this present review, the progress in regeneration, in vitro propagation, chimeral segregation, callus and protoplast culture, embryo rescue, in vitro germination, and genetic transformation of roses were discussed and the impact of biotechnology on rose breeding was evaluated.

Keywords: Rose, tissue culture, genetic transformation, breeding.

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V.M. Kulkarni

Papers

Isolation and regeneration of rose bud callus protoplasts (Protoplasts sp. cv. soraya)