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Salvia splendens is a widely used ornamental bedding plant; however, the limited propagation method has decreased its quality and yield. Through years of selection, we have obtained a new variety of S. splendens with weak apical dominance and named it as ‘Cailinghong’. To establish an effective method for regeneration of S. splendens ‘Cailinghong’, different explants, including leaves, receptacles, petioles, stem nodes, and stem segments were used for adventitious bud induction. Next, various combinations of plant growth regulators (PGRs) were selected for bud and root induction, which were assessed by adventitious bud initiation rate and proliferation rate, as well as root induction rate. Meanwhile, the survival rate of transplanted plantlets was also calculated. As a result, stem nodes were found easy to be induced to form buds, and the optimum medium component was 1/2 Murashige and Skoog (MS) medium supplemented with 0.45 mM naphthalene acetic acid (NAA), 8.88 mM 6-benzylaminopurine (6-BA), and 2.46 mM 3-indolebutyric acid (IBA) for plantlets induction, whereas 1/4 MS medium supplemented with 2.23 mM NAA for root induction. Furthermore, the survival rate of transplanted plantlets was up to 80%, and all regenerated plantlets were normal in phenotype. Therefore, cultured in 1/2 MSmedium with combined PGRs, whole plantlet of S. splendens ‘Cailinghong’ could be regenerated directly from stem node. Among all the numerous ornamental bedding plants, due to its bright colors and long florescence, S. splendens belonging to Labiatae family, is most widely planted in gardens, flowerbeds, and by roadsides (Soundararajan et al., 2013). With the development of research and technology, the breeding of S. splendens has been largely improved (Liu et al., 2012), and many new varieties have been cultivated, such as S. splendens ‘Zhongchuanhong’, a tall variety breed (Hui et al., 2004), and S. splendens ‘Shenzhouhong’, a variety produced through space breeding (Fu et al., 2009). It has been reported that, wild type S. splendens as well as its new varieties need to remove the terminal bud three or four times to meet the product requirements (Zhi et al., 2013). Through years of selection, we have found a new variety with weak apical dominance and named it as ‘Cailinghong’. Its original lateral branch develops into the first lateral branch during vegetative growth phase, which further develops into multilevel lateral branches. ‘Cailinghong’ grows and develops without the need of manual removal of the terminal bud during its whole life. At last, it forms a naturally low and spherical type plant with multilevel lateral branches and blossom exuberant, which is more suitable for landscaping. As far as we know, there is no other variety of S. splendens having such powerful branching ability as ‘Cailinghong’ does. In addition, related report about its tissue culture remains rare. However, there are still many problems in the development of S. splendens, such as limited colors, serious pests, and diseases, which will inevitably lead to a decrease in yield and a decline in flower quality. What is more, S. splendens shows higher requirements on temperature, which adds difficulties on its reproduction (Lai et al., 2001). Thus, it is of great importance to investigate proper rapid propagation technique of S. splendens. Recently, an indirect regeneration method that uses the callus of S. splendens has been reported. Unfortunately, the regeneration cycle is comparatively long.Moreover, under in vitro conditions, because of long-term exposure to PGRs and chemicals, the callus cells are very sensitive (Maruthi Rao et al., 2012), and they can exhibit abnormal morphological characteristics and somaclonal variation. Direct regeneration by organogenesis is an alternative method, referring to the culture of different tissues in vitro such as buds, leaves, stems, and roots, to develop new shoots (Mall on et al., 2011). In all, it is an effective method of mass propagation of plants, while there is no need for callus as an intermediate stage. In this study, explants (receptacle, leaf, petiole, stem segment, and stem node) were selected to establish an effective method for regeneration of S. splendens ‘Cailinghong’. Then the effects of PGRs on adventitious bud induction and shoot regeneration were evaluated. The results may provide clues for further application of biotechnology on regeneration of S. splendens ‘Cailinghong’. Materials and Methods Plant materials. S. splendens ‘Cailinghong’ was purchased from Zhejiang Hongyue Flower Co. Ltd (Hangzhou, Zhejiang, China). Explants (receptacles, leaves, petioles, stem nodes, and stem segments) were obtained from 2to 4month-old mature ‘Cailinghong’ grown in the greenhouse (22 ± 2 C with humidity of 60%). Selection of explants. Receptacles were obtained from flower bud whose height was about 1 cm. Leaves, petioles, and stems were collected from tender branches and leaves of the collected stem nodes were removed. Then explants of leaf were cut into small pieces of 5 mm · 5 mm, and explants of petiole, stem node, and stem segment were cut into small pieces of 5 to 9 mm. In this way, each stem node only contained two axillary buds. Explants were surface sterilized by 75% (v/v) ethanol for 30 s, then by 1% (v/v) sodium hypochlorite for 1 min, and finally rinsed with sterile distilled water for three times (3 min each time). Three explants of receptacle, leaf, petiole, stem node, or stem segment were planted in each glass vessel (five glasses for each type of explant). All explants were cultured in glass vessels that contained 1/2 MS basal culture medium (Murashige and Skoog, 1962) without PGRs. The initiation rate was observed and calculated: Initiation rate = (No. of initiated explants/No. of inoculated explants) · 100%. Induction of adventitious bud. By calculating the initiation rates of different types of explants, the explant with the highest initiation rate was chosen as the optimal explant. Then the overall experiment of all the possible combinations was designed and the sterilized optimal explants were placed on 1/2 MS medium (pH 5.8) supplemented with different concentrations and combinations of NAA (0, 0.45, or 2.23 mM), 6-BA (4.44, 8.88, or 22.20 mM), and IBA (0, 2.46, or 4.92 mM) to determine the optimum medium for adventitious bud induction (Soundararajan et al., 2013). The concentrations of 13.32 and 17.76 mM 6-BA were excluded because Received for publication 4 Sept. 2014. Accepted for publication 7 Jan. 2015. This study was supported by the Beijing Municipal Commission of Education—Science and Technology Project (PXM2014_014207_1), Scientific Research Quality Improvement Project of the Beijing University of Agriculture (GZL-2013007), and Youth Science Foundation of the Beijing University of Agriculture (2117516004). We wish to express our heartfelt thanks to Fenghe (Shanghai) Information Technology Co., Ltd. Their ideas and help gave a valuable added dimension to our research. The authors have declared that no competing interests exist. To whom reprint requests should be addressed; e-mail kefengliu67@hotmail.com. HORTSCIENCE VOL. 50(6) JUNE 2015 869 GROWTH REGULATORS

Adventitious Bud Induction and Plant Regeneration from Stem Nodes of Salvia splendens ‘Cailinghong’