论文部分内容阅读
Abstract In order to explore the best storage temperature and storage period of dragon fruit seeds, with the constant-humidity germplasm cold storage of Guangxi Academy of Agricultural Sciences as a platform and storage temperature as an environmental factor, the germination characteristics of dragon fruit seeds and seedling growth characteristics under low temperature storage (8, 4, -1, -10 ℃) and room temperature storage (25-30 ℃) over time were explored. The results showed that with the extension of the storage time, the seed germination rates, germination index, healthy seedling rates, seedling heights and fresh weights of 10 seedlings of the three dragon fruit varieties showed basically the same trends, showing a downward trend. After 367 d of cold storage, the germination rate decreased from 100% to 92.33%-98.00%, and the room temperature treatments decreased to 54.00%-86.30%; and the healthy seedling rates of cold-stored seeds decreased from 96.33%-98.00% to 77.33%-97.3%, and the values of the room temperature treatments dropped to 12.00%-68.33%. The germination rates, germination index, healthy seedling rates and seedling heights of the three dragon fruit varieties stored at low temperatures were basically significantly higher than those of the room temperature treatments within 367 d of storage. The germination rates of the three varieties stored at 8 ℃ had no significant differences from other low temperature treatments, but the germination index was significantly lower than other low temperature treatments at 367 d. It showed that low temperature storage at -10-8 ℃ could significantly prolong the vigor and storage time of dragon fruit seeds. Considering cost and other factors, it is better to store dragon fruit seeds in a dry environment at 4 ℃. The seeds stored at 4 ℃ for 1 year showed a germination rate maintained over 90% and a healthy seedling rate over 85%.
Key words Low temperature storage; Dragon fruit; Seed germination; Seedlings; Genebank
Received: July 13, 2020 Accepted: September 14, 2020
Supported by Basic Operating Expenses of Guangxi Academy of Agricultural Sciences (GNK 2015YT98, GNK 2020YM21, GNK 2020YM20).
Yihao XING (1991-), male, P. R. China, assistant research fellow, master, devoted to research about storage and management of crop germplasm resources.
*Corresponding author. Dongqiang WEN(1971-), male, P. R. China, senior engineer, master, devoted to research about the management, maintenance and technical renovation of the germplasm bank equipment and facilities. E-mail: wdqab@163.com. Dragon fruit (Hylocereus spp.) is a unique tropical berry fruit with small but many seeds, shaped like sesame, evenly distributed in the flesh. Each fruit has 4 000-6 000 seeds. The 1 000-seed weight of dragon fruit is 1.0-1.8 g, and the seeds have thin coat, can absorb water easily, and can germinate under suitable conditions after being taken from mature fruit generally. However, during the seedling process of seeds (including conventional seeds and hybrids), it was discovered that after being harvested, dragon fruit seeds stored at room temperature and humidity for 0.5 to 1 year will severely lose their viability, which leads to extremely low germination rate, which is not conducive to sowing of the same batch of seeds at different times within a year. In addition, dragon fruit is mostly produced in southern subtropical or tropical regions, where there is more rain in spring and high air humidity and high temperature in summer, which lead to strong seed respiration, strong metabolism and high consumption of nutrients, which is extremely unfavorable to the storage of dragon fruit seeds. Therefore, it is very important to explore suitable storage conditions for dragon fruit seeds to maintain their viability and extend the life of the seeds.
Storage temperature and seed water content are the most important factors affecting seed viability during storage[1]. Storing seeds in a low-temperature genebank is considered to be an effective long-term method of preserving plant germplasm resources, and is also the key to preserving biodiversity[2]. The current researches on dragon fruit seeds mainly focus on seed germination characteristics[3-5], culture conditions[6-7], stress response[8-12] and chemical mutagenesis[13], but rarely on their storage characteristics. Therefore, in this study, with the constant-humidity germplasm cold storage of Guangxi Academy of Agricultural Sciences as a platform and storage temperature as an environmental factor, the germination characteristics of dragon fruit seeds and seedling growth and change law at different storage temperatures were explored, aiming to clarify the best storage conditions and storage period and to provide references for the preservation and utilization of dragon fruit seeds.
Materials and Methods
Materials
The seeds of Meilong 1, Hongmeigui 5 red-skin and red-flesh and Vietnam Bairou red-skin and white-flesh dragon fruit varieties were selected as materials. Their 1 000-seed weights were 1.47, 1.53, and 1.30 g, respectively. Seed collection and processing
The middle flesh of mature fruit of the test dragon fruit was put in a nylon mesh bag with small apertures, and rubbed to clean the flesh tissue outside the seed. The seeds were rinsed with clean water and dried in a cool place to get the test seeds. Impurities and seeds that were not full, shriveled or broken were removed before storage.
Experimental design
The experiment was carried out on September 26, 2018. The seeds were stored at four low temperatures, namely -10, -1, 4, 8 ℃, in the germplasm genebank of Guangxi Academy of Agricultural Sciences (humidity stabilized at 35%-45%), and the seeds in a desiccator at room temperature (25-30 ℃) were used as a control and stored continuously for 367 d. During the storage period, the seeds of each storage temperature were regularly taken out for germination test, and 0 d was used as the pre-storage control. Each treatment was repeated 3 times, and each replicate included 100 seeds.
Germination test
According to the International Rules for Seed Testing, seeds of each treatment were sterilized with 10% sodium hypochlorite and placed in sterile water for 6 h. Two layers of filter paper were placed at the bottom of a petri dish with a diameter of 9 cm, and the filter paper was moistened with a small amount of ddH2O. Dragon fruit seeds were evenly placed on the filter paper for the germination test, and then placed in an artificial climate box at 25 ℃ for constant temperature cultivation, with a photoperiod of 12 h, and a light intensity of 25 000 Lx. An appropriate amount of sterile water was added every day to keep the filter paper moist.
Statistical analysis of data
During the germination period, the starting date of germination, the number of seeds that germinated normally (the seeds showed white radicle with a length greater than the seed radius) and the number of normal and healthy seedlings in each treatment were observed and recorded. The test was terminated when no more germination occurred for 3 consecutive days. The germination rate was counted, and the germination index was calculated. After 24 d of culture, the percentage of healthy seedlings was counted, and 10 seedlings were randomly selected in each replicate to determine the height (the distance from the base to the cotyledon, that is, the height of the hypocotyl) and the total fresh weight of the 10 seedlings. The calculation equations were as follows: Germination rate (%) = Number of germinated seeds/Number of tested seeds×100, Germination index = ΣGt/Dt (Gt is the number of seeds germinated on day t, and Dt is the corresponding number of germination days), healthy seedling rate (%) = Number of healthy seedlings/Number of tested seeds ×100. Excel 2010 statistical software was used to sort out the data and draw graphs. Single factor and two factor analysis of variance used SPSS22.0, and multiple comparisons used Duncan method. The germination rate and healthy seedling rate data represented by percentage (%) were transformed by arc sine to perform variance analysis.
Results and Analysis
Effects on the germination characteristics of dragon fruit seeds during storage
Different storage temperatures had significant effects on the germination rate and germination index of the three dragon fruit varieties (P<0.05). The germination rates of Meilong 1 and Vietnam Bairou treated at room temperature were significantly lower than that of low temperature treatments at 124 d of storage (P<0.05); the germination rates of the three varieties had hardly any significant differences in different low temperature treatments at 367 d of storage (P>0.05), and with the decrease of storage temperature, the germination rates did not show a certain trend of change. However, in the same storage period, the germination index of different low temperature treatments showed different degrees of variety difference. The germination index of Hongmeigui 5 had no significant differences between different low temperatures at 367 d of storage (P>0.05); and the germination index of Vietnam Bairou stored at 8 ℃ was significantly lower than other low temperature treatments at 301 d of storage (P<0.05), while that of Meilong 1 was significantly lower than other low temperature treatments until it was stored for 367 d. The germination index of Meilong 1, Hongmeigui 5 and Vietnam Bairou treated at room temperature was significantly lower than other low temperature treatments at 34, 184, and 124 d (P<0.05), respectively. In addition, when the three dragon fruit varieties were stored for 367 d, their germination index showed an increasing trend to varying degrees with the decrease of storage temperature (Table 1 and Table 2).
Different storage days had significant effects on the germination rate and germination index of the three dragon fruit varieties (P<0.05). With the extension of storage days, the seed germination rate and germination index of the three dragon fruit varieties decreased, and the trends of change were basically the same, but the significance and extent of the decline showed variety differences under the same storage temperature. Compared with 0 d (before storage), the germination rate and germination index of the three dragon fruit varieties at room temperature were significantly lower than other low temperature treatments. When stored for 367 d, Meilong 1 showed the smallest germination rate and germination index at room temperature, of 54.00% and 6.56, respectively, followed by Vietnam Bairou (61.00% and 11.20) and Hongmeigui 5 (86.33% and 25.13). The germination rate and germination index of the seeds stored at each low temperature were maintained over 92.33% and 27.65, respectively, after 367 d of storage (Table 1 and Table 2). It shows that low-temperature storage can significantly slow down the decline of seed germination vigor. Effects on the growth of dragon fruit seedlings during storage
Different storage temperatures had significant effects on the healthy seedling rate, seedling height and fresh weight of 10 seedlings of the three dragon fruit varieties (P<0.05). The healthy seedling rates and seedling heights of the three dragon fruit varieties stored at room temperature were all lower than those of the low temperature treatments within 367 d, and the fresh weights of 10 seedlings of the three dragon fruit varieties stored at room temperature were also lower than the low temperature treatments at most storage time. The fresh weight of 10 seedlings of Vietnam Bairou stored at room temperature was always lower than low temperature treatments within 367 d, while Meilong 1 and Hongmeigui 5 were lower than low temperature treatments until 184 d. The effects of low temperature storage on healthy seedling rate, seedling height and fresh weight of 10 seedlings of dragon fruit were different between varieties. From the perspective of seedling growth index, the effects from high to low ranked as Meilong 1>Hongmeigui 5>Vietnam Bairou overall. In the low temperature treatments, the 4 ℃ treatments of the three dragon fruit varieties exhibited higher healthy seedling rates, seedling heights and fresh weights of 10 seedlings.
The room temperature is 25-30 ℃; different uppercase letters indicate significant differences between treatments in the same column (P<0.05), and different lowercase letters indicate significant differences between treatments in the same row (P<0.05). The same in Table 2.
Different storage time had significant effects on the healthy seedling rates, seedling heights and 10-seedling fresh weights of the three dragon fruit varieties (P<0.05). With the extension of storage time, the healthy seedling rates, seedling heights and fresh weights of 10 seedlings of the three dragon fruit varieties had basically the same changing trends, all showing a downward trend, but there were variety differences in the speed and significance of the decline. The healthy seedling rate, seedling height and fresh weight of 10 seedlings of Vietnam Bairou were the lowest when stored at room temperature for 367 d, which were 12.00%, 6.17 mm and 0.18 g, respectively, followed by Meilong 1 (40.33%, 9.20 mm and 0.26 g) and Hongmeigui 5 (68.33%, 9.88 mm and 0.31 g). The decreases in healthy seedling rate, seedling height and fresh weight of 10 seedlings of seeds stored at low temperatures were significantly lower than those at room temperature after 184 d of storage. Among them, the healthy seedling rates of Meilong 1 and Hongmeigui 5 stored at 4 ℃ decreased to small extents within 367 d of storage, and the differences were not significant (P>0.05) (Fig. 1 and Table 3). Conclusions and Discussion
Seed vigor is closely related to storage conditions. To maintain high vigor, proper storage conditions are the key. During the storage of seeds, water content and storage environmental conditions are one of the important factors affecting seed viability[14-16], among which storage temperature and humidity are important factors in maintaining seed viability during storage[17]. However, seeds of different species have great differences in their own water content and storage environment temperature and humidity requirements. It has been reported that the vitality of some plant seeds will decline rapidly under the conditions of high temperature and high humidity, and low temperature and dry conditions will delay the senescence of the seeds, can maintain high viability for a long time, and help the long-term preservation of the seeds[18-20]. The evaluation of seed vigor usually uses germination characteristics such as germination rate and germination index as indicators for testing[21-23], and the determination of seedling growth indicators, including seedling height (hypocotyl) and fresh weight, can also be used for seed quality and vigor evaluation indexes[24]. In this study, the three varieties of dragon fruit seeds had higher germination rates and healthy seedling rates under different low temperature storage conditions. With the extension of the storage time, their decline rates were significantly slower than those of the seeds stored at room temperature, and the values still remained above 92.33% and 77.33% after storage for 367 d, respectively, which were much higher than the room temperature seeds. It indicated that low temperature storage at -10-8 ℃ could significantly delay the decline of seed vigor and prolong seed life. It also further confirmed that it is feasible to maintain the vigor of dragon fruit seeds by reducing the storage temperature. When the three dragon fruit varieties were stored for 367 d, the germination index showed different degrees of increasing trend with the decrease of storage temperature, which showed that with the extension of storage period, lower storage temperature was beneficial to the maintenance of dragon fruit seed vigor and helped extend seed life. The storage characteristics of dragon fruit seeds are consistent with some crop seeds[15, 19, 25-28]. In addition, this study showed that the germination rates of the three varieties stored at 8 ℃ had no significant differences from other low temperature treatments, but the germination index was significantly lower than other low temperature treatments at 367 d, which indicated that although 8 ℃ had little effect on the germination rate of seeds, the ability to delay the decline of seed vigor was significantly reduced with the extension of the storage period, and the differences in the germination characteristics between the 4, -1, and -10 ℃ treatments were relatively small. The physiological characteristics of the variety itself is also another important factor affecting seed viability[29-30]. Seeds with dormancy characteristics are usually more storable at room temperature[31-34], while seeds without dormancy characteristics are lower[26, 35]. The three dragon fruit varieties selected in this study could germinate on the second day after sowing during the 367 d storage period, and no obvious dormancy phenomenon was found. Moreover, dragon fruit seeds are smaller and have less internal nutrients, which may be one of the reasons why dragon fruit easily loses seed vigor at room temperature. In this study, the germination rates of the three dragon fruit varieties did not differ greatly among the low temperatures, but they showed obvious differences in the germination speed (germination index) and seedling growth (healthy seedling rate, seedling height and 10-seedling fresh weight) among varieties. Among them, the seed vigor of Meilong 1 under low temperature storage conditions decreased slowly with the extension of storage time, and showed higher storability. Whether it is related to the strong cold tolerance of this variety needs further research. In terms of seedling growth indicators, the healthy seedling rate, seedling height and fresh weight of 10 seedlings of Hongmeigui 5 were the highest when stored at room temperature for 367 d, showing strong seedling growth. Whether it is related to the larger seeds of this variety (its 1 000-seed weight is 1.53 g) containing more nutrients needs further study.
In this study, the germination characteristics and seedling growth indexes of the three dragon fruit varieties stored at 4 ℃ were relatively good, and had smaller differences from the -1 and -10 ℃ storage treatments, and the seeds stored for 1 year showed a germination rate maintained over 90% and a healthy seedling rate over 85%. Considering the practical feasibility, economic cost and other factors, it is better to dry and store dragon fruit seeds at 4 ℃. The impact of storage humidity as an important factor in environmental conditions on the vitality of dragon fruit seeds during storage will be the focus of the next step.
References
[1] MAKEEN AM, NORMAH MN, DUSSERT S, et al. Seed moisture characteristics in relation to total lipid content of five Citrus taxa using an equilibrium dehydration protocol[J]. Seed Science and Technology, 2006, 34(2): 453-464.
[2] GONZLEZ-BENITO ME, ALBERT MJ, IRIONDO JM, et al. Seed germination of four thyme species after short-term storage at low temperatures at several moisture contents[J]. Seed Science and Technology, 2004, 32(1): 247-254. [3] WANG B, ZHENG W, CAI YQ, et al. Study on germination characteristics of dragon fruit seeds[J]. Seed, 2007, 26(5): 16-19. (in Chinese)
[4] WU L, LI JX, QIN X, et al. Study on germination characteristics of different flesh color dragon fruit seeds[J]. Seed, 2016, 35(3): 74-77. (in Chinese)
[5] ZHAO CX, WANG XY, LI HB, et al. Preliminary study on germination characteristics of Hylocereus undatus seed[J]. Seed, 2005, 24(8): 37-40. (in Chinese)
[6] YANG GR, TANG AJ. Analysis on germination characteristics of light-requiring pitahaya (Hylocereus undulates) seeds[J]. Seed, 2017, 36(10): 80-83. (in Chinese)
[7] HU ZY, LIANG GD, HUANG HS, et al. Exogenous GA3 presoaking treatment to dragon fruit seeds for improving germination[J]. Genomics and Applied Biology, 2010, 29(3): 529-533. (in Chinese)
[8] DENG RJ. Study on physiological response of dragon fruit to low temperature stress and screening of cold resistance mutants by in vitro mutation[D]. Ya’an: Sichuan Agricultural University, 2015. (in Chinese)
[9] LIU HG, DUAN RT, SHA SC, et al. Effect of PEG-6000 treatment on seed germination of dragon fruit[J]. Acta Agriculturae Jiangxi, 2010, 22(11): 50-52. (in Chinese)
[10] WANG B, ZHENG W, LI XZ. The effect of seed soaking with aluminum on seed germination of Hylocereus polyrhizus (Weber) Britt. & Rose[J]. Seed, 2006, 25(4): 38-40. (in Chinese)
[11] ZHANG HJ, YU CC, YU J, et al. Effects of cadmium on seed germination and antioxidant enzyme activity of cotyledon in dragon fruit[J]. Molecular Plant Breeding, 2016, 14(11): 3172-3176. (in Chinese)
[12] ZHENG W, WANG B, CAI YQ, et al. Effect of NaCl stress on seed germination of dragon fruit[J]. Seed, 2008, 27(1): 82-85. (in Chinese)
[13] ZHANG BX, WEN CP. Effects of EMS treatment on the seed germination of dragon fruit[J]. Journal of Mountain Agriculture and Biology, 2013, 32(3): 194-197. (in Chinese)
[14] HAN YZ, WU L, ZENG B, et al. Effects of storage temperature and seed moisture content on seed vigor of tall fescue[J]. Seed, 2011, 30(6): 41-44. (in Chinese)
[15] ZHU P, KONG LQ, LI G, et al. Effect of moisture content on physiological characteristics of Elymus sibiricus seed under different storage temperature conditions[J]. Acta Prataculturae Sinica, 2011, 20(6): 101-108. (in Chinese)
[16] TANG HH, ZHAO Q, XU DX, et al. Effects of four Acacia seeds vitality under different storage conditions[J]. Forestry and Environmental Science, 2019, 35(5): 68-73. (in Chinese) [17] TIAN Q, XIN X, LU XX, et al. Research progress on plant seed aging and mitochondria[J]. Journal of Plant Genetic Resources, 2012, 13(2): 283-287. (in Chinese)
[18] DU M, MU YH, YUAN SW, et al. Analysis on storage factors affecting rice seed vigor[J]. Soil and Crop, 2018, 7(4): 419-422. (in Chinese)
[19] GAO GZ, WU XM, LYU XD, et al. Genotype differences of seed viability in rapeseed during storage at different temperature[J]. Chinese Journal of Oil Crop Sciences, 2010, 32(4): 495-499. (in Chinese)
[20] GUO KT. Study on seedling vigor of field crop at different storage condition[J]. Journal of Shaoguan University: natural science, 2003, 24(6): 85-87. (in Chinese)
[21] LUO SW, WU JH, ZHONG L. Effect of temperature on the germination characteristics of Shuicheng tall fescue seeds[J]. Agriculture and Technology, 2020, 40(2): 14-15. (in Chinese)
[22] MA Z, WANG AB. Effects of different temperature gradients on seed germination of four turfgrass species[J]. Modern Horticulture, 2020(5): 31-32. (in Chinese)
[23] QIN AL, GUO QS, MA FQ, et al. Effects of temperature, light and water conditions on seed germination of Thuja sutchuenensis Franch[J]. Seed, 2020, 39(2): 15-20. (in Chinese)
[24] FILHO CPH, GONELI ALD, MASETTO TE, et al. The effect of drying temperatures and storage of seeds on the growth of soybean seedlings[J]. Journal of Seed Science, 2016, 38(4): 287-295.
[25] DA SILVA GH, TOLEDO MZ, TEIXEIRA RN, et al. Influence of the storage environment on the physiological quality of millet seeds (Pennisetum glaucum (L.) R.Br.)[J]. Journal of Seed Science, 2019, 41(3): 286-292.
[26] LUO Q, GUO XL, WANG AY, et al. Effect of different storage conditions on seed germination rate of Angelica pubescens seed[J]. Modern Chinese Medicine, 2019, 21(8): 1080-1083. (in Chinese)
[27] PIAO J, SHAO TY, LYU LS. Effects of different storage and treatment methods on germination rate of Bupleurum seeds in Changbai Mountain[J]. Journal of Agricultural Science Yanbian University, 2007, 29(1): 27-29. (in Chinese)
[28] DECRUSE SW, SEENI S. Seed cryopreservation is a suitable storage procedure for a range of Piper species[J]. Seed Science and Technology, 2003, 31(1): 213-217.
[29] BALESEVIC-TUBICS, TATICM, DORDEVICV, et al. Seed viability of oil crops depending on storage conditions[J]. Helia, 2010, 33(52): 153-160.
[30] ARIYARATHNA RIS, WEERASENA SL, BENERAGAMA CK. Sustenance of seed germination potential of two okra (Abelmoschus esculentus L. Moench)varieties under different environmental conditions in Sri Lanka[J].Tropical Agricultural Research, 2020, 31(1): 21-30. [31] FU N, SONG H, WANG SJ, et al. Research progress on seed dormancy and breaking methods[J]. Journal of Anhui Agricultural Sciences, 2018, 46(24): 10-12. (in Chinese)
[32] NIU YQ, XU HB, ZHANG YX, et al. Studies on molecular mechanism of plant seed dormancy[J]. Fujian Journal of Agricultural Sciences, 2017, 32(10): 1156-1164. (in Chinese)
[33] YAO LJ, ZHANG KL, XIONG ZM, et al. Research advances in seed morphophysiological dormancy[J]. Chinese Journal of Ecology, 2019, 38(1): 247-255. (in Chinese)
[34] RODRIGUEZ MV, BODRONE MP, CASTELLARI MP, et al. Effect of storage temperature on dormancy release of sunflower (Helianthus annuus) achenes[J].Seed Science Research, 2018, 28(2): 101-111.
[35] YANG P, XIAN MZ, HU LY, et al. Effects of low-temperature on seed germination and seedling development of rapeseed[J]. Journal of Huazhong Agricultural University, 2015, 34(6): 7-13. (in Chinese)
Editor: Yingzhi GUANG Proofreader: Xinxiu ZHU
Key words Low temperature storage; Dragon fruit; Seed germination; Seedlings; Genebank
Received: July 13, 2020 Accepted: September 14, 2020
Supported by Basic Operating Expenses of Guangxi Academy of Agricultural Sciences (GNK 2015YT98, GNK 2020YM21, GNK 2020YM20).
Yihao XING (1991-), male, P. R. China, assistant research fellow, master, devoted to research about storage and management of crop germplasm resources.
*Corresponding author. Dongqiang WEN(1971-), male, P. R. China, senior engineer, master, devoted to research about the management, maintenance and technical renovation of the germplasm bank equipment and facilities. E-mail: wdqab@163.com. Dragon fruit (Hylocereus spp.) is a unique tropical berry fruit with small but many seeds, shaped like sesame, evenly distributed in the flesh. Each fruit has 4 000-6 000 seeds. The 1 000-seed weight of dragon fruit is 1.0-1.8 g, and the seeds have thin coat, can absorb water easily, and can germinate under suitable conditions after being taken from mature fruit generally. However, during the seedling process of seeds (including conventional seeds and hybrids), it was discovered that after being harvested, dragon fruit seeds stored at room temperature and humidity for 0.5 to 1 year will severely lose their viability, which leads to extremely low germination rate, which is not conducive to sowing of the same batch of seeds at different times within a year. In addition, dragon fruit is mostly produced in southern subtropical or tropical regions, where there is more rain in spring and high air humidity and high temperature in summer, which lead to strong seed respiration, strong metabolism and high consumption of nutrients, which is extremely unfavorable to the storage of dragon fruit seeds. Therefore, it is very important to explore suitable storage conditions for dragon fruit seeds to maintain their viability and extend the life of the seeds.
Storage temperature and seed water content are the most important factors affecting seed viability during storage[1]. Storing seeds in a low-temperature genebank is considered to be an effective long-term method of preserving plant germplasm resources, and is also the key to preserving biodiversity[2]. The current researches on dragon fruit seeds mainly focus on seed germination characteristics[3-5], culture conditions[6-7], stress response[8-12] and chemical mutagenesis[13], but rarely on their storage characteristics. Therefore, in this study, with the constant-humidity germplasm cold storage of Guangxi Academy of Agricultural Sciences as a platform and storage temperature as an environmental factor, the germination characteristics of dragon fruit seeds and seedling growth and change law at different storage temperatures were explored, aiming to clarify the best storage conditions and storage period and to provide references for the preservation and utilization of dragon fruit seeds.
Materials and Methods
Materials
The seeds of Meilong 1, Hongmeigui 5 red-skin and red-flesh and Vietnam Bairou red-skin and white-flesh dragon fruit varieties were selected as materials. Their 1 000-seed weights were 1.47, 1.53, and 1.30 g, respectively. Seed collection and processing
The middle flesh of mature fruit of the test dragon fruit was put in a nylon mesh bag with small apertures, and rubbed to clean the flesh tissue outside the seed. The seeds were rinsed with clean water and dried in a cool place to get the test seeds. Impurities and seeds that were not full, shriveled or broken were removed before storage.
Experimental design
The experiment was carried out on September 26, 2018. The seeds were stored at four low temperatures, namely -10, -1, 4, 8 ℃, in the germplasm genebank of Guangxi Academy of Agricultural Sciences (humidity stabilized at 35%-45%), and the seeds in a desiccator at room temperature (25-30 ℃) were used as a control and stored continuously for 367 d. During the storage period, the seeds of each storage temperature were regularly taken out for germination test, and 0 d was used as the pre-storage control. Each treatment was repeated 3 times, and each replicate included 100 seeds.
Germination test
According to the International Rules for Seed Testing, seeds of each treatment were sterilized with 10% sodium hypochlorite and placed in sterile water for 6 h. Two layers of filter paper were placed at the bottom of a petri dish with a diameter of 9 cm, and the filter paper was moistened with a small amount of ddH2O. Dragon fruit seeds were evenly placed on the filter paper for the germination test, and then placed in an artificial climate box at 25 ℃ for constant temperature cultivation, with a photoperiod of 12 h, and a light intensity of 25 000 Lx. An appropriate amount of sterile water was added every day to keep the filter paper moist.
Statistical analysis of data
During the germination period, the starting date of germination, the number of seeds that germinated normally (the seeds showed white radicle with a length greater than the seed radius) and the number of normal and healthy seedlings in each treatment were observed and recorded. The test was terminated when no more germination occurred for 3 consecutive days. The germination rate was counted, and the germination index was calculated. After 24 d of culture, the percentage of healthy seedlings was counted, and 10 seedlings were randomly selected in each replicate to determine the height (the distance from the base to the cotyledon, that is, the height of the hypocotyl) and the total fresh weight of the 10 seedlings. The calculation equations were as follows: Germination rate (%) = Number of germinated seeds/Number of tested seeds×100, Germination index = ΣGt/Dt (Gt is the number of seeds germinated on day t, and Dt is the corresponding number of germination days), healthy seedling rate (%) = Number of healthy seedlings/Number of tested seeds ×100. Excel 2010 statistical software was used to sort out the data and draw graphs. Single factor and two factor analysis of variance used SPSS22.0, and multiple comparisons used Duncan method. The germination rate and healthy seedling rate data represented by percentage (%) were transformed by arc sine to perform variance analysis.
Results and Analysis
Effects on the germination characteristics of dragon fruit seeds during storage
Different storage temperatures had significant effects on the germination rate and germination index of the three dragon fruit varieties (P<0.05). The germination rates of Meilong 1 and Vietnam Bairou treated at room temperature were significantly lower than that of low temperature treatments at 124 d of storage (P<0.05); the germination rates of the three varieties had hardly any significant differences in different low temperature treatments at 367 d of storage (P>0.05), and with the decrease of storage temperature, the germination rates did not show a certain trend of change. However, in the same storage period, the germination index of different low temperature treatments showed different degrees of variety difference. The germination index of Hongmeigui 5 had no significant differences between different low temperatures at 367 d of storage (P>0.05); and the germination index of Vietnam Bairou stored at 8 ℃ was significantly lower than other low temperature treatments at 301 d of storage (P<0.05), while that of Meilong 1 was significantly lower than other low temperature treatments until it was stored for 367 d. The germination index of Meilong 1, Hongmeigui 5 and Vietnam Bairou treated at room temperature was significantly lower than other low temperature treatments at 34, 184, and 124 d (P<0.05), respectively. In addition, when the three dragon fruit varieties were stored for 367 d, their germination index showed an increasing trend to varying degrees with the decrease of storage temperature (Table 1 and Table 2).
Different storage days had significant effects on the germination rate and germination index of the three dragon fruit varieties (P<0.05). With the extension of storage days, the seed germination rate and germination index of the three dragon fruit varieties decreased, and the trends of change were basically the same, but the significance and extent of the decline showed variety differences under the same storage temperature. Compared with 0 d (before storage), the germination rate and germination index of the three dragon fruit varieties at room temperature were significantly lower than other low temperature treatments. When stored for 367 d, Meilong 1 showed the smallest germination rate and germination index at room temperature, of 54.00% and 6.56, respectively, followed by Vietnam Bairou (61.00% and 11.20) and Hongmeigui 5 (86.33% and 25.13). The germination rate and germination index of the seeds stored at each low temperature were maintained over 92.33% and 27.65, respectively, after 367 d of storage (Table 1 and Table 2). It shows that low-temperature storage can significantly slow down the decline of seed germination vigor. Effects on the growth of dragon fruit seedlings during storage
Different storage temperatures had significant effects on the healthy seedling rate, seedling height and fresh weight of 10 seedlings of the three dragon fruit varieties (P<0.05). The healthy seedling rates and seedling heights of the three dragon fruit varieties stored at room temperature were all lower than those of the low temperature treatments within 367 d, and the fresh weights of 10 seedlings of the three dragon fruit varieties stored at room temperature were also lower than the low temperature treatments at most storage time. The fresh weight of 10 seedlings of Vietnam Bairou stored at room temperature was always lower than low temperature treatments within 367 d, while Meilong 1 and Hongmeigui 5 were lower than low temperature treatments until 184 d. The effects of low temperature storage on healthy seedling rate, seedling height and fresh weight of 10 seedlings of dragon fruit were different between varieties. From the perspective of seedling growth index, the effects from high to low ranked as Meilong 1>Hongmeigui 5>Vietnam Bairou overall. In the low temperature treatments, the 4 ℃ treatments of the three dragon fruit varieties exhibited higher healthy seedling rates, seedling heights and fresh weights of 10 seedlings.
The room temperature is 25-30 ℃; different uppercase letters indicate significant differences between treatments in the same column (P<0.05), and different lowercase letters indicate significant differences between treatments in the same row (P<0.05). The same in Table 2.
Different storage time had significant effects on the healthy seedling rates, seedling heights and 10-seedling fresh weights of the three dragon fruit varieties (P<0.05). With the extension of storage time, the healthy seedling rates, seedling heights and fresh weights of 10 seedlings of the three dragon fruit varieties had basically the same changing trends, all showing a downward trend, but there were variety differences in the speed and significance of the decline. The healthy seedling rate, seedling height and fresh weight of 10 seedlings of Vietnam Bairou were the lowest when stored at room temperature for 367 d, which were 12.00%, 6.17 mm and 0.18 g, respectively, followed by Meilong 1 (40.33%, 9.20 mm and 0.26 g) and Hongmeigui 5 (68.33%, 9.88 mm and 0.31 g). The decreases in healthy seedling rate, seedling height and fresh weight of 10 seedlings of seeds stored at low temperatures were significantly lower than those at room temperature after 184 d of storage. Among them, the healthy seedling rates of Meilong 1 and Hongmeigui 5 stored at 4 ℃ decreased to small extents within 367 d of storage, and the differences were not significant (P>0.05) (Fig. 1 and Table 3). Conclusions and Discussion
Seed vigor is closely related to storage conditions. To maintain high vigor, proper storage conditions are the key. During the storage of seeds, water content and storage environmental conditions are one of the important factors affecting seed viability[14-16], among which storage temperature and humidity are important factors in maintaining seed viability during storage[17]. However, seeds of different species have great differences in their own water content and storage environment temperature and humidity requirements. It has been reported that the vitality of some plant seeds will decline rapidly under the conditions of high temperature and high humidity, and low temperature and dry conditions will delay the senescence of the seeds, can maintain high viability for a long time, and help the long-term preservation of the seeds[18-20]. The evaluation of seed vigor usually uses germination characteristics such as germination rate and germination index as indicators for testing[21-23], and the determination of seedling growth indicators, including seedling height (hypocotyl) and fresh weight, can also be used for seed quality and vigor evaluation indexes[24]. In this study, the three varieties of dragon fruit seeds had higher germination rates and healthy seedling rates under different low temperature storage conditions. With the extension of the storage time, their decline rates were significantly slower than those of the seeds stored at room temperature, and the values still remained above 92.33% and 77.33% after storage for 367 d, respectively, which were much higher than the room temperature seeds. It indicated that low temperature storage at -10-8 ℃ could significantly delay the decline of seed vigor and prolong seed life. It also further confirmed that it is feasible to maintain the vigor of dragon fruit seeds by reducing the storage temperature. When the three dragon fruit varieties were stored for 367 d, the germination index showed different degrees of increasing trend with the decrease of storage temperature, which showed that with the extension of storage period, lower storage temperature was beneficial to the maintenance of dragon fruit seed vigor and helped extend seed life. The storage characteristics of dragon fruit seeds are consistent with some crop seeds[15, 19, 25-28]. In addition, this study showed that the germination rates of the three varieties stored at 8 ℃ had no significant differences from other low temperature treatments, but the germination index was significantly lower than other low temperature treatments at 367 d, which indicated that although 8 ℃ had little effect on the germination rate of seeds, the ability to delay the decline of seed vigor was significantly reduced with the extension of the storage period, and the differences in the germination characteristics between the 4, -1, and -10 ℃ treatments were relatively small. The physiological characteristics of the variety itself is also another important factor affecting seed viability[29-30]. Seeds with dormancy characteristics are usually more storable at room temperature[31-34], while seeds without dormancy characteristics are lower[26, 35]. The three dragon fruit varieties selected in this study could germinate on the second day after sowing during the 367 d storage period, and no obvious dormancy phenomenon was found. Moreover, dragon fruit seeds are smaller and have less internal nutrients, which may be one of the reasons why dragon fruit easily loses seed vigor at room temperature. In this study, the germination rates of the three dragon fruit varieties did not differ greatly among the low temperatures, but they showed obvious differences in the germination speed (germination index) and seedling growth (healthy seedling rate, seedling height and 10-seedling fresh weight) among varieties. Among them, the seed vigor of Meilong 1 under low temperature storage conditions decreased slowly with the extension of storage time, and showed higher storability. Whether it is related to the strong cold tolerance of this variety needs further research. In terms of seedling growth indicators, the healthy seedling rate, seedling height and fresh weight of 10 seedlings of Hongmeigui 5 were the highest when stored at room temperature for 367 d, showing strong seedling growth. Whether it is related to the larger seeds of this variety (its 1 000-seed weight is 1.53 g) containing more nutrients needs further study.
In this study, the germination characteristics and seedling growth indexes of the three dragon fruit varieties stored at 4 ℃ were relatively good, and had smaller differences from the -1 and -10 ℃ storage treatments, and the seeds stored for 1 year showed a germination rate maintained over 90% and a healthy seedling rate over 85%. Considering the practical feasibility, economic cost and other factors, it is better to dry and store dragon fruit seeds at 4 ℃. The impact of storage humidity as an important factor in environmental conditions on the vitality of dragon fruit seeds during storage will be the focus of the next step.
References
[1] MAKEEN AM, NORMAH MN, DUSSERT S, et al. Seed moisture characteristics in relation to total lipid content of five Citrus taxa using an equilibrium dehydration protocol[J]. Seed Science and Technology, 2006, 34(2): 453-464.
[2] GONZLEZ-BENITO ME, ALBERT MJ, IRIONDO JM, et al. Seed germination of four thyme species after short-term storage at low temperatures at several moisture contents[J]. Seed Science and Technology, 2004, 32(1): 247-254. [3] WANG B, ZHENG W, CAI YQ, et al. Study on germination characteristics of dragon fruit seeds[J]. Seed, 2007, 26(5): 16-19. (in Chinese)
[4] WU L, LI JX, QIN X, et al. Study on germination characteristics of different flesh color dragon fruit seeds[J]. Seed, 2016, 35(3): 74-77. (in Chinese)
[5] ZHAO CX, WANG XY, LI HB, et al. Preliminary study on germination characteristics of Hylocereus undatus seed[J]. Seed, 2005, 24(8): 37-40. (in Chinese)
[6] YANG GR, TANG AJ. Analysis on germination characteristics of light-requiring pitahaya (Hylocereus undulates) seeds[J]. Seed, 2017, 36(10): 80-83. (in Chinese)
[7] HU ZY, LIANG GD, HUANG HS, et al. Exogenous GA3 presoaking treatment to dragon fruit seeds for improving germination[J]. Genomics and Applied Biology, 2010, 29(3): 529-533. (in Chinese)
[8] DENG RJ. Study on physiological response of dragon fruit to low temperature stress and screening of cold resistance mutants by in vitro mutation[D]. Ya’an: Sichuan Agricultural University, 2015. (in Chinese)
[9] LIU HG, DUAN RT, SHA SC, et al. Effect of PEG-6000 treatment on seed germination of dragon fruit[J]. Acta Agriculturae Jiangxi, 2010, 22(11): 50-52. (in Chinese)
[10] WANG B, ZHENG W, LI XZ. The effect of seed soaking with aluminum on seed germination of Hylocereus polyrhizus (Weber) Britt. & Rose[J]. Seed, 2006, 25(4): 38-40. (in Chinese)
[11] ZHANG HJ, YU CC, YU J, et al. Effects of cadmium on seed germination and antioxidant enzyme activity of cotyledon in dragon fruit[J]. Molecular Plant Breeding, 2016, 14(11): 3172-3176. (in Chinese)
[12] ZHENG W, WANG B, CAI YQ, et al. Effect of NaCl stress on seed germination of dragon fruit[J]. Seed, 2008, 27(1): 82-85. (in Chinese)
[13] ZHANG BX, WEN CP. Effects of EMS treatment on the seed germination of dragon fruit[J]. Journal of Mountain Agriculture and Biology, 2013, 32(3): 194-197. (in Chinese)
[14] HAN YZ, WU L, ZENG B, et al. Effects of storage temperature and seed moisture content on seed vigor of tall fescue[J]. Seed, 2011, 30(6): 41-44. (in Chinese)
[15] ZHU P, KONG LQ, LI G, et al. Effect of moisture content on physiological characteristics of Elymus sibiricus seed under different storage temperature conditions[J]. Acta Prataculturae Sinica, 2011, 20(6): 101-108. (in Chinese)
[16] TANG HH, ZHAO Q, XU DX, et al. Effects of four Acacia seeds vitality under different storage conditions[J]. Forestry and Environmental Science, 2019, 35(5): 68-73. (in Chinese) [17] TIAN Q, XIN X, LU XX, et al. Research progress on plant seed aging and mitochondria[J]. Journal of Plant Genetic Resources, 2012, 13(2): 283-287. (in Chinese)
[18] DU M, MU YH, YUAN SW, et al. Analysis on storage factors affecting rice seed vigor[J]. Soil and Crop, 2018, 7(4): 419-422. (in Chinese)
[19] GAO GZ, WU XM, LYU XD, et al. Genotype differences of seed viability in rapeseed during storage at different temperature[J]. Chinese Journal of Oil Crop Sciences, 2010, 32(4): 495-499. (in Chinese)
[20] GUO KT. Study on seedling vigor of field crop at different storage condition[J]. Journal of Shaoguan University: natural science, 2003, 24(6): 85-87. (in Chinese)
[21] LUO SW, WU JH, ZHONG L. Effect of temperature on the germination characteristics of Shuicheng tall fescue seeds[J]. Agriculture and Technology, 2020, 40(2): 14-15. (in Chinese)
[22] MA Z, WANG AB. Effects of different temperature gradients on seed germination of four turfgrass species[J]. Modern Horticulture, 2020(5): 31-32. (in Chinese)
[23] QIN AL, GUO QS, MA FQ, et al. Effects of temperature, light and water conditions on seed germination of Thuja sutchuenensis Franch[J]. Seed, 2020, 39(2): 15-20. (in Chinese)
[24] FILHO CPH, GONELI ALD, MASETTO TE, et al. The effect of drying temperatures and storage of seeds on the growth of soybean seedlings[J]. Journal of Seed Science, 2016, 38(4): 287-295.
[25] DA SILVA GH, TOLEDO MZ, TEIXEIRA RN, et al. Influence of the storage environment on the physiological quality of millet seeds (Pennisetum glaucum (L.) R.Br.)[J]. Journal of Seed Science, 2019, 41(3): 286-292.
[26] LUO Q, GUO XL, WANG AY, et al. Effect of different storage conditions on seed germination rate of Angelica pubescens seed[J]. Modern Chinese Medicine, 2019, 21(8): 1080-1083. (in Chinese)
[27] PIAO J, SHAO TY, LYU LS. Effects of different storage and treatment methods on germination rate of Bupleurum seeds in Changbai Mountain[J]. Journal of Agricultural Science Yanbian University, 2007, 29(1): 27-29. (in Chinese)
[28] DECRUSE SW, SEENI S. Seed cryopreservation is a suitable storage procedure for a range of Piper species[J]. Seed Science and Technology, 2003, 31(1): 213-217.
[29] BALESEVIC-TUBICS, TATICM, DORDEVICV, et al. Seed viability of oil crops depending on storage conditions[J]. Helia, 2010, 33(52): 153-160.
[30] ARIYARATHNA RIS, WEERASENA SL, BENERAGAMA CK. Sustenance of seed germination potential of two okra (Abelmoschus esculentus L. Moench)varieties under different environmental conditions in Sri Lanka[J].Tropical Agricultural Research, 2020, 31(1): 21-30. [31] FU N, SONG H, WANG SJ, et al. Research progress on seed dormancy and breaking methods[J]. Journal of Anhui Agricultural Sciences, 2018, 46(24): 10-12. (in Chinese)
[32] NIU YQ, XU HB, ZHANG YX, et al. Studies on molecular mechanism of plant seed dormancy[J]. Fujian Journal of Agricultural Sciences, 2017, 32(10): 1156-1164. (in Chinese)
[33] YAO LJ, ZHANG KL, XIONG ZM, et al. Research advances in seed morphophysiological dormancy[J]. Chinese Journal of Ecology, 2019, 38(1): 247-255. (in Chinese)
[34] RODRIGUEZ MV, BODRONE MP, CASTELLARI MP, et al. Effect of storage temperature on dormancy release of sunflower (Helianthus annuus) achenes[J].Seed Science Research, 2018, 28(2): 101-111.
[35] YANG P, XIAN MZ, HU LY, et al. Effects of low-temperature on seed germination and seedling development of rapeseed[J]. Journal of Huazhong Agricultural University, 2015, 34(6): 7-13. (in Chinese)
Editor: Yingzhi GUANG Proofreader: Xinxiu ZHU