Lonicera L., commonly known as honeysuckle, is a genus of woody plants and belongs to the family Caprifoliaceae. There are 180 recognized species of deciduous or evergreen, bushy, candent, twining or creeping shrubs distributed in Asia, Europe, and North America (Huxley, 1994). Flowers are epigynous, paired, and axillary or in usually six-flowered whorls on terminal spikes with different colors and a pronounced fragrance. Thus, honeysuckles have been widely grown as ornamental plants (Georges et al., 1992; Huxley, 1994; Karhu, 1997). Because of their different growth habitats and great adaptability, a few species such as L. japonica Thunb. (Schierenbeck, 2004) and L. maackii (Rupr.) Herder. (Gorchov and Trisel, 2003) are also considered invasive species in some regions.
Dried flowers and buds of honeysuckles are called “Jinyinhua” in Chinese or flos lonicerae, which is a famous herb of traditional Chinese medicine. Extracts of flos lonicerae have a wide range of benefits to human health, including anti-inflammation, bacteriostasis, antivirus, antioxidant, and hematischesis (Lee et al., 1998; Palacios et al., 2002; Peng et al., 2005). Flos lonicerae contains a series of water-soluble phenolic compounds with 3-O-ceffeoyl-D-quinic acid or chlorogenic acid (CGA) being a major ingredient. In addition to having antipyretic properties, CGA has been reported to suppress N-nitrosating reactions and inhibit hepatic glucose 6-phosphatase, which may be a significant factor in the abnormal diabetic state (Arion et al., 1997; Yang et al., 2004). CGA has been studied in L. confuse DC., L. dasystyla Rehder, L. hypoglauca Miq. (Guo and Wan, 2005; Li et al., 2003), L. japonica (Lee et al., 1998), and L. tatarica L. (Palacios et al., 2002).
Another species, L. macranthoides Hand.-Mazz., native to China (Xu and Wang, 1988), was found to contain much higher concentrations of CGA (4.0% to 4.5%) than L. japonica (2.2% to 2.5%) (Zhou and Tong, 2003). A survey of genetic resources of Lonicera in southwest Hunan province, China, in 1997 to 1998 resulted in the identification of a mutant from a L. macranthoides population that was heavily infected with powdery mildew disease (Microsphaera lonicerae DC.). The mutant (Fig. 1A), in contrast to its wild type (Fig. 1B), resists the powdery mildew and possesses abundant flowers that never open. Wang et al. (2004) documented that the mutant had an average of 30 nonopened flowers per axillary and the flower longevity ranged from 15 to 25 d compared with 5 to 7 d of the wild-type plants. The mutant was named ‘Jincuilei’. CGA concentration in the nonopened flowers of ‘Jincuilei’ was higher, up to 6.0% compared with 4.3% in its wild-type plants. This mutant, however, does not produce seeds and propagation by cutting or grafting has only a ≈30% survival rate (Wang et al., 2004), which significantly hampers its commercial production as a medicinal plant.
In vitro culture has been shown to be an efficient method of propagating medicinal plants (Debnath et al., 2006; Rout et al., 2000). Research on micropropagation of ‘Jincuilei’ started in 2000. A method of eliminating contaminants from leaf explants was established (Wang et al., 2005). B5 medium (Gamborg et al., 1968), Murashige and Skoog (MS) medium (Murashige and Skoog, 1962), woody plant medium (Lloyd and McCown, 1980), and White basal medium (White, 1963) were tested for culture of leaf explants. However, a simple and efficient protocol for rapid regeneration of ‘Jincuilei’ was not well developed. The objectives of this study were to establish an efficient procedure for inducing callus and adventitious shoot formation of ‘Jincuilei’ and determine if the regenerated plants maintained the nonopened flower characteristic and high level of CGA.
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