Studies on the Bulb Development and Its Physiological Mechanisms in Lilium Oriental Hybrids Y.P. Xia, H.J. Zheng and C.H. Huang Department of Horticulture College of Agriculture and Biotechnology Zhejiang Univeresty, Hangzhou China Keywords: Lilium Oriental hybrids, bulb development, mountainous region, carbohydrate Abstract Based on the growth observation of lily bulbs (Lilium Oriental hybrids, Acapulco, Marco Polo, Siberia, Sobonne and Tiber ), we found the mountainous condition was suitable for the bulb development. Total 288,2 bulbs (12-14 cm, 14-16 cm and 16-18 cm in circumference) of Oriental lily were harvested from 41, imported bulblets (6/9 cm), which were grown in the mountainous region (Jinyun, Zhejiang Province) having an elevation of 88 m. The average ratio for harvested flowering bulbs (>12 cm) and big bulbs (16-18 cm) was 71.7% and 15.4%, respectively. The concentrations of soluble sugar, sucrose and starch in the bulb scales, as well as leaf photosynthesis were investigated during the whole year cultivation. It was found that the fresh weight and circumference of harvested bulbs have the positive correlation between vegetative growth in lily. The bud removal improved the process of bulb development. Accumulation and transportation of carbohydrates in the bulbs also increased significantly from the bud-removal. The average concentrations of starch and total soluble sugar were 293 mg g FW -1 and 9.2 mg g FW -1 in the harvested bulbs, respectively. It was indicated that the quality of mountainous reproduction bulbs was similar to that of the commercial bulbs from Holland. INTRODUCTION Although the origin of 47 species and 18 varieties for genus Lilium, Chinese domestic production of commercial lily bulbs has only taken place for about 1 years (Xia and Gao, 21; Zhao and Zhao, 21). Regions including southwest, northeast, and east of China are the main areas for the production of lily bulbs, while Yunnan province and the mountainous region in Zhejiang province are particularly suitable for Oriental lily (Wei, 23; Xia et al., 23). It has been suggested that early removal of lily flowers should enhance lily bulb yield (Blaney and Roberts, 1967; Wang and Breen, 1986). Assimilate from a given leaf is partitioned in different patterns during plant development due to the changing carbon demand of receiving organs (Matsuo and Mizuno, 1974; Wang and Breen, 1986). Several authors have looked for growth and development of the Easter lily, and the carbohydrate changes were studied especially related during cold treatment or regenerated in vitro (Fujimoto et al., 1989; Miller and Langhans, 1989; Shin et al., 22). It is unclear to the accumulation and regulators of carbohydrate in bulb development for Oriental lily. The objectives of this study were to describe the domestic reproduction of lily bulbs in mountainous region of Zhejiang province, and to compare the physiological mechanisms with the imported commercial bulbs for achieving the domestication of lily bulbs in China. MATERIALS AND METHODS Total 41, bulblets (6-9 cm in circumference) were imported from the Netherlands, and cultivars included Acapulco, Marco Polo, Siberia, Sorbonne and Tiber. All the bulblets were planted in Dayang Mountain (Jinyun county, Zhejiang Proc. IX th Intl. Symp. on Flower Bulbs Eds.: H. Okubo, W.B. Miller and G.A. Chastagner Acta Hort. 673, ISHS 25 91
province) having an elevation of 88 m during mid April 23. The soil in the mountainous region is sandy acid red-yellow soil. Flower buds were removed in late July, and the bulbs were harvested at the end of November 23. The data statistics for plant growth and bulb development were the average of 2 samples. The second scale from outmost bulbs ( Sorbonne and Siberia ) were used for carbohydrate analysis. Fresh samples were placed in 8% ethanol to separate an ethanol-soluble fraction containing soluble sugars. The ethanol-soluble supernatant was taken after centrifugation for 15 min at 8, x g. The precipitates were suspended in 52% HClO 4 to extract the starch after removing the ethanol soluble sugar residues. Anthrone reaction was used for carbohydrate determination, including the concentrations of soluble sugar and starch. The concentration of reducing sugar was determined using 3,5-Dinitrosalicylic acid, and the sucrose content was the balance of total soluble sugar minus the reducing sugar. An Infrared Rays CO 2 Analysis System 255-MK 3 (ADC, Hoddesdon, England) was used for determination of net photosynthetic rate. RESULTS AND DISCUSSION Six growth stages from April 22 to November 15 for sampling were separated in this paper, represented by A-F (Table 1). Lily Bulbs Reproduction Even though the plant growth and the blooming capability of lily were normal when grown in natural condition of Hangzhou, bulb reproduction was limited due to the high temperatures that occurred from late May. The climatic condition in mountainous region of Dayang is similar to the Netherlands and the Mediterranean region, which was relatively cool for lily growth and bulb development during the summer (Table 2). A total of 288,2 bulbs (12-14 cm, 14-16 cm and 16-18 cm in circumference) of Oriental lily ( Acapulco, Marco Polo, Siberia, Sorbonne and Tiber ) were harvested from 41, imported bulblets (6/9 cm). About 68.8-9.4% of bulblets (6-9 cm) became flowering bulbs (>12 cm) after one year s growth for five popular cultivars of Oriental lily (Table 3). The average ratio for harvested flowering bulbs was 71.7%, and the ratios of big size (16-18cm in circumference) are from 9.3% to 3.% for these five cultivars. Plant Growth and Bulb Development The fresh weight of bulbs increased after growth stage B, while the bulb size significantly increased after bud removal (Fig. 1). About 3 days after bud removal, increases in plant height began to slow, which resulted in leaves that were more tolerant of the summer time growing conditions (Fig. 2). Compared with the two most important cultivars Siberia and Sorbonne, the sizes of harvested bulbs showed different efficacy for bulb reproduction. About 16.6% of the bulblets became 16/18 cm bulbs for Sorbonne, while with Siberia only 9.3% were of the bigger size (Table 4). This indicated that in the mountainous region in Zhejiang, conditions were more suitable for the growth of bulblets of Sorbonne than Siberia. The amount of leaves on the stems of plants was related to the size of harvested bulbs. The fresh weight of bulb and size of harvested bulb were significantly higher in plants with more than 18 leaves (Table 5). Behavior of Carbohydrates in Lily Bulbs The concentrations of soluble sugar and sucrose in bulbs decreased after planting, while starch content increased during all growth stages (Fig. 3). The average concentrations of starch and total soluble sugar in the harvested bulbs were 293 mg g FW -1 and 9.2 mg g FW -1, respectively. Starch content increased dramatically after bud removal (Fig. 4). This indicates that starch accumulates rapidly after blooming, and that bud removal improved the transportation of carbohydrates to the developing bulbs. Sucrose has been shown to be the main soluble carbohydrate associated with bulb 92
development (Gude and Verbruggen, 2; Shin et al., 22). More than half the total soluble sugars detected at the various growth stages was sucrose (Table 1). The starch content was also compared with commercial lily bulbs and Chinese traditional lily (Lilium davidii var. unicolor) (Sun, H.M. 23, Ph.D. dissertation) (Table 6). The results showed there was no differences with the starch contents, but the concentrations of soluble sugar and sucrose were lower than that found in commercial bulbs. Net Photosynthesis and Protein Analysis Analysis of net photosynthesis indicted the saturation level occurred when PFD was 74 µmol m -2 s -1 (Fig. 5). It was clear that the natural levels of sunlight are suitable for lily growth. Comparison of SDS-PAG electrophoresis of Oriental lily showed a marked difference in the scale and the leaf, but no obvious difference was found within various growth stages in the scales (Fig. 6). Two coarse bands in the scale suggested that there was some stable fractions of protein associated with bulb development. CONCLUSIONS The mountainous regions in Zhejiang province were suitable for plant growth and bulb development of Oriental lilies. About 7% of imported bulblets became commercial flowering bulbs after one year s growth. Flower bud removal benefited plant growth and improved bulb reproduction. Accumulation and transportation of carbohydrates increased after effloresce. Both bulb size and starch content of the reproduction bulbs grown in the mountainous region were similar to levels found in commercial bulbs. ACKNOWLEDGEMENTS Thanks to Jiaxing BIYUN Garden Company for financial assistance. Literature Cited Blaney, L.T. and Roberts, A.N. 1967. Growth and development of the Easter lily bulb. Proc. Amer. Soc. Hortic. Sci. 89:643-6. Fujimoto, S., Kitahara, K. and Hamaya, R. 1989. Studies on Easter lily starch. Bull. Fac. Agric. Kagoshima Univ. 39:121-13. Gude, H. and Verbruggen, J. 2. Physiological markers for lily bulb maturity. Acta Hort. 517:343-3. Matsuo, T. and Mizuno, T. 1974. Changes in the amount of two kinds of reserve glucose-containing polysaccharides during germination of the Easter lily bulb. Plant Cell Physiol. 15:555-558. Miller, W.B. and Langhans, R.W. 1989. Carbohydrate changes of Easter lilies during growth in normal and reduced irradiance environments. J. Amer. Soc. Hort. Sci. 114:31-315. Shin, K.S., Chakrabarty, D. and Paek, K.Y. 22. Sprouting rate, change of carbohydrate contents and related during cold treatment of lily bulblets regenerated in vitro. Scientia Hort. 96:195-24. Wang, Y.T. and Breen, P.J. 1986. Partitioning of 14 C-assimilate in Easter lily as affected by growth stage and flower removal. Scientia Hort. 29:273-281. Wei, Z.X. 23. Precedence domestic production of lily bulbs in Yunnan. China Flowers & Hort. 22:16-18. Xia, Y.P. and Gao, X.C. 21. The development and problems of domestic production for bulb flowers in China. Advances in Floriculture of China. China Agriculture Press. p.216-223. Xia, Y.P., Gao, X.C. and Zheng, H.J. 23. Discussion about the development of domestic production for lily bulbs in China. China Flowers & Hort. 22:11-13. Zhao, X.Y. and Zhao, W.Y. 21. Remark on the development and strategy for lily industry in China. Advances in Floriculture of China. China Agriculture Press. 93
p.129-132. Tables Table 1. Growth stages at sampling and comparison of the ratios for sucrose content to total soluble sugar content. Date 4/22 6/15 7/15 8/8 9/25 11/15 Growth period Planting stage Growth stage Visible bud stage After bud removal Bulb developed Withered stage Symbol A B C D E F Ratio (%) 96.1 76.5 64.1 44.7 65.3 75.2 Date: month/day Table 2. Comparison of the climatic conditions in Dayang Mountains, Hangzhou, the Mediterranean region, and the Netherlands. Areas Average temperature in January ( C) Average temperature in July ( C) Annual rainfall (mm) Netherlands 2. 16. 6~7 Mediterranean 1 1. 23. 7 Hangzhou 4.2 29.5 1,226 Dayang Mountain 2 2.3 24.5 1,789 1 Mediterranean region (4 N, E) and the Netherlands ( N, 5 E): average data from climate maps 2 Dayang Mountain elevation is 88 m Table 3. Yield of flowering bulbs (>12 cm) and big size (16/18 cm) bulbs from imported bulblets (6/9 cm). Cultivar Planted Harvested bulbs bulblets Total amount (>12 cm) Ratio (%) Big size (16/18 cm) Ratio (%) Acapulco 11, 9,94 9.4 3,3 3. Marco Polo 74,8 52,98 69.6 17,17 23. Siberia 196, 135,385 68.9 18,37 9.3 Sorbonne 76, 56,178 73.9 12,64 16.6 Tiber 45, 34,42 76.5 1,3 23. Total amount 41, 288,21 71.7 61,83 15.4 94
Table 4. Comparison of various sizes of harvested bulbs from bulblets of Oriental lily Siberia and Sorbonne. Harvested bulbs Siberia (6/9 cm) (Total 196,) Sorbonne (6/9 cm) (Total 76,) 16/18 cm Total amount Ratio (%) 18,37 9.3 12,64 16.6 14/16 cm Total amount Ratio (%) 25,37 12.9 17,318 22.8 12/14 cm Total amount 91,645 26,22 Ratio (%) 46.6 34.5 Table 5. Influence of leaf number on the plant growth and bulb development Sorbonne. Leaves Plant height (cm) Fresh weight of bulb (g) Bulb size (cm) 9/25 1/11 11/25 9/25 1/11 11/25 9/25 1/11 11/25 >2 52A 62A 55A.6A 59.7A 85.4A 16.2A 16.6A 19.1A 18~2 46B 48B 45B 52.2A 68.6A 45.9AB 15.8A 16.2A 18.2A 15~17 45B 4B 41B 34.1B 4.9B 45.AB 14.4AB 14.7AB 15.4B <15 38B 39B 4B 28.1B 33.7B 38.B 12.4B 13.5B 14.5B Significant response at P <.1 (**) Table 6. Comparison of carbohydrates contents between the reproduction bulbs and commercial bulbs in lily. Bulb samples Starch content (mg g FW -1 ) Soluble sugar (mg g FW -1 ) Sucrose content (mg g FW -1 ) Regenerated bulb 293 9.2 6.9 Commercial bulb 28~3 3~4 1~2 Lilium davidii var. unicolor 2~29 7~9 ~7 95
Figures Fresh weight Bulb size Fresh weight of bulb(g) 6 4 3 2 1 18 15 12 9 6 3 Bulb size(cm) A B C D E F Growth period Fig. 1. Changes in Lilium Oriental hybrid Sorbonne bulb development in the mountainous region at various growth periods. 6 Bud removal Normal Plant height (cm) 4 3 2 1 2 4 6 8 1 12 Days after bud removal Fig. 2. Comparison of Lilium Oriental hybrid Siberia plant growth following removal of flower buds. 96
Soluble sugar Sucrose Starch Content of soluble sugar and sucrose (mg gfw -i ) 7 6 4 3 2 1 A B C D E F 3 3 2 2 1 1 Concentration of Starch (mg gfw -i ) Growth period Fig. 3. Changes in the concentrations of soluble sugar, sucrose and starch in lily bulbs during various growth stages in Sorbonne. Starch content mg.gfw -1 2 Bud removal 2 Normal 1 1 3 6 9 12 1 Days after planting Fig. 4. Changes in the starch content in bulbs after planting and flower bud removal of Siberia. 97
7 Pn (µmolco2m-2s-1) 6 5 4 3 Upside leaves 2 Middle leaves 1-1 1 2 3 4 6 7 8 9 1-2 Fig. 5. Light response curves of photosynthesis in lily leaf of Siberia. PFD (µmolm-2s-1) 1 2 3 4 5 6 Fig. 6. Comparison of SDS-PAG electrophoresis in lily scale and leaf tissue. 1) Visible bud stage; 2) 1d after effloresce; 3) 4d after effloresce; 4) Harvest stage; 5) Upside leaves; 6) Underside leaves (Lilium Oriental hybrid Siberia ). 98