Physiological Changes and Related Genes Expression Analysis of Sorghum (Sorghum bicolor) Seedlings Under Low Temperature Stress
SHAO Wen-Jing, ZHANG Jin-Jie, GE Sheng-Nan, WEI Yu-Lei, LI Jia-Xin, WU Geng-Jin, LIU Xin-Yu, HE Lin, WEI Jin-Peng*, XU Jing-Yu*
Key Laboratory of Modern Agricultural Cultivation and Crop Germplasm Improvement of Heilongjiang Province, Heilongjiang Engineering Technology Research Center for Crop Straw Utilization, Heilongjiang Province Cultivating Collaborative Innovation Center for The Beidahuang Modern Agricultural Industry Technology, National Coarse Cereal Engineering Research Center, College of Agriculture, Heilongjiang Bayi Agricultural University, Daqing 163319, China
Abstract：Low temperature is one of the critical environmental stress affecting the growth and development of sorghum (Sorghum bicolor) in the cold region. In order to study the physiological changes of sorghum seedlings leaves and the expression of related genes under low temperature stress, the cold-resistance variety 'P61' and the cold-sensitive variety 'H21' were selected as the test materials, which were subjected to low-temperature treatment at 4 ℃ for 0, 1, 3, 5 and 7 d, respectively. The results showed that the chlorophyll fluorescence parameters (Fv/Fm) and relative chlorophyll content (SPAD) decreased significantly with the extension of the low-temperature treatment time, whereas the decrease was not obvious in the cold-tolerant variety 'P61'; the expression of genes related to photosynthesis were down-regulated with the prolonged stress time, and the expression level was higher in resistant varieties; the activity of superoxide dismutase (SOD) and peroxidase (POD) was elevated after 5~7 d treatment. At 7 d, the SOD activity of 'H21' decreased by 3.3%, the SOD activity of 'P61' increased by 15.2%, and the POD activity of 'H21' and 'P61' increased by 7.4% and 9%, respectively. The content of malondialdehyde (MDA) and proline (Pro) increased. The MDA content of 'P61' and 'H21' increased by 36.5% and 1.7%, respectively, the content of Pro increased by 69% and 230.5%. A set of 8 SbSODs genes were screened from transcriptome database, the number of SbSODs up-regulated at low temperature for 3 d were larger than those treated for 1 d, and the expression of SbSOD5 was up-regulated in the low-temperature treatment of both varieties for 1 d and 3 d; 7 sorghum PODs genes were screened, and the most of the genes (5 out of 7) were up-regulated in both varieties when treated for 1 d, Sobic.009G055100 was up-regulated in both varieties after 1 d and 3 d treatment; a total of 11 SbLOXs were screened, and a number of up-regulated SbLOXs (lipoxygenase) were observed at 1 d time point, and SbLOX9 was up-regulated in both varieties under 1 d and 3 d treatments; 2 sorghum P5CS genes were selected, of which Sobic.003G356000 was significantly up-regulated at different time points in two varieties, and a high expression level (Log2FC>6 ) was detected in 'P61' when treated for 3 d. This study provides a reference for further research on the physiological regulation of plants under low temperature stress and the transcriptional regulation of related genes.
[1] 蔡志欢, 张桂莲. 2018. 水稻低温冷害研究进展[J]. 作物研究, 32(3): 249-255. (Cai Z H, Zhang G L.2018. Research progress of low temperature and chilling injury of rice[J]. Journal of Crop Research, 32(3): 249-255.) [2] 曹梦可. 2016. 玉米耐寒能力分级及低温对苗期玉米光合生理机制的影响[D]. 硕士学位论文,吉林农业大学, 导师: 吴春胜, pp. 29-42. (Cao M K.2016. Screening of cold resistance of different maize cultivars and effect of low temperature on photosynthesis and physiology mechanism of maize during seeding stage[D]. Thesis for M.S., Jilin Agriculture University, Supervisor: Wu C S, pp. 29-42.) [3] 陈浩维. 2018. 干旱和低温对玫瑰生理生化影响及4 ℃低温下SOD基因表达分析[D]. 硕士学位论文, 昆明理工大学, 导师: 文锦芬, pp. 33-41. (Chen H W.2018. Study on physiological and biochemical response to drought stress and low temperature in rose and SOD gene expression under 4 ℃ low temperature[D]. Thesis for M.S., Kunming University of Science and Technology University, Supervisor: Wen J F, pp: 33-41.) [4] 陈梅, 唐运来. 2012. 低温胁迫对玉米幼苗叶片叶绿素荧光参数的影响[J]. 内蒙古农业大学学报(自然科学版), 33(3): 20-24. (Chen M, Tang Y L.2012. Effects of low temperature stress on chlorophyll fluorescence characteristics of com seedlings[J]. Journal of Inner Mongolia Agricultural University (Natural Science Edition), 33(3): 20-24.) [5] 陈明辉, 张志录, 程世平, 等. 2016. 低温胁迫对不同品种果蔗膜脂过氧化及渗透调节物质的影响[J]. 作物杂志, 175(6): 53-57. (Cheng M H, Zhang Z L, Cheng S P, et al.2016. Effects of low temperature stress on lipid peroxidation and osmotic regulation substances of chewing cane seedlings[J]. Journal of Crop Magazine, 175(6): 53-57.) [6] 古丽江·许库尔汗, 孙雅丽, 阿依古丽·铁木儿, 等. 2019. 低温胁迫对c枝条渗透物质含量、膜质过氧化及保护酶活性的影响[J]. 新疆农业科学, 56(4): 685-695. (Gu L J, Sun Y L, A Y G L, et al.2019. Effects of low temperature stress on the content of penetration substance, membrane peroxidation and protective enzyme activity in Ribes rubrum[J]. Journal of Xinjian Agricultural Science, 56(4): 685-695.) [7] 郭慧, 李树杏, 孙平勇等. 2020. 水稻苗期耐冷性差异的转录组分析[J]. 分子植物育种, 18(06): 1731-1739. (Guo H, Li S X, Sun P Y, 2020. Transcriptomes analysis of the cold tolerance difference of rice seedlings[J]. Molecular Plant Breeding, 18(06): 1731-1739.) [8] 郝平安, 梁芳, 张燕,等. 2018. 低温胁迫对蝴蝶兰光合及生理特性的影响[J]. 热带作物学报, 39(10): 1955-1962. (Hao P A, Liang F, Zhang Y, et al.2018. Effects of cold stress on the photosynthetic and physiological characteristics of phalaenopsis[J]. Journal of Tropical Crops, 39(10): 1955-1962.) [9] 黄丽芳, 刘建汀, 王彬, 等. 2018. 西葫芦过氧化物酶POD1基因的分离及表达分析[J]. 福建农业学报, 33(09): 943-949. (Isolation and expression of peroxidase POD1 gene from Cucurbita pepo[J]. Fujian Journal of Agriculture, 33(09): 943-949) [10] 刘瑞媛, 董喜存, 梁开平, 等. 2018. 不同品种甜高粱苗期耐寒性分析研究[J]. 中国农学通报, 34(31): 19-26. (Liu R Y, Dong X C, Liang K P, et al.2018. Chilling tolerance of different sweet sorghum varieties at seedling stage[J]. Chinese Agricultural Science Bulletin, 34(31): 19-26. ) [11] 聂书明, 杜中平, 王丽慧, 等. 2016. 低温弱光对辣椒叶片光合特性和生理特性的影响[J]. 西南农业学报, 29(10): 2319-2323. (Nie S M, Du Z P, Wang L H, et al .2016. Effect of photosynthesis characteristics and physiological characteristics of pepper seedlings under low temperature and weak light stress[J]. Southweast China Journal of Agricultural Sciences, 29(10): 2319-2323. ) [12] 潘铁夫, 冯绍印, 郑秀梅,等. 1980. 东北地区高梁低温冷害及其防御[J]. 吉林农业科学, 41(4): 29-38. (Pan T F, Feng S Y, Zheng X M, et al.1980. Sorghum low temperature cold damage and its defense in northeast China[J]. Journal of Jilin Agricultural Sciences, 41(4): 29-38.) [13] 平璐. 2016. 紫花苜蓿Cu/Zn-SOD基因在矮牵牛中的表达及其耐冷性分析[D]. 硕士学位论文. 吉林农业大学, 导师: 顾德峰, pp. 37-44. (Ping L, 2016. Over-expression of Ms Cu/Zn-SOD gene in Petunia hybrida and the cold tolerance analysis of transgenic plants[D]. Thesis for M.S., Jilin Agriculture University, Supervisor: Gu D W, pp. 37-44.) [14] 史红梅, 张海燕, 杨彬, 等. 2015.低温胁迫对高粱幼苗MDA含量、SOD和POD活性的影响[J]. 中国农学通报, 31(18): 74-79. (Shi H M, Zhang H Y, Yang B, et al.2015. Effects of low temperature stress on the content of MDA, SOD and POD activity in sorghum seedlings[J]. Journal of China Agricultural Communications, 31(18): 74-79.) [15] 舒必超, 杨勇, 刘雪勇, 等. 2018. 低温胁迫对狗牙根生理及基因表达的影响[J]. 草业学报, 27(11): 106-119. (Shu B C, Yang Y, Liu X Y, et al.2018. Effect of low temperature stress on physiology and gene expression in Bermuda grass[J]. Acta Prataculturae Sinica, 27(11): 106-119.) [16] 王瑞, 马凤鸣, 李彩凤, 等. 2008. 低温胁迫对玉米幼苗脯氨酸、丙二醛含量及电导率的影响[J]. 东北农业大学学报, 159(5): 20-23. (Wang R, Ma F M, Li C F, et al.2008. Effect of low temperature stress on proline, malondialdehyde contents and electric conductivity of maize seedling[J]. Journal of Northeast Agricultural University, 159(5): 20-23.) [17] 吴建慧, 杨玲, 孙国荣. 2004. 低温胁迫下玉米幼苗叶片活性氧的产生及保护酶活性的变化[J]. 植物研究, 2004(04): 456-459. (Wu J H, Yang L, Sun G R.Generation of activated oxygen and change of cell defense enzyme activity in leaves of maize seedling under the stress of low temperature[J]. Bulletin of Botanical Research, 2004(04): 456-459.) [18] 吴锦程, 吴毕莎, 黄审剑, 等. 2015. 枇杷幼果PLD和LOX对低温胁迫的响应[J]. 植物科学学报, 33(2): 203-209. (Wu J C, Wu B S, Huang S J, et al.2015. Phospholipase D and liposygenase of young loquat fruits in response to low temperature stress[J]. Journal of Plant Science. 33(2): 203-209.) [19] 吴雪霞, 朱月林, 朱为民, 等. 2006. 外源一氧化氮对NaCl胁迫下番茄幼苗生理影响[J]. 中国农业科学, 15(3): 575-581. (Wu X X, Zhu Y L, Zhu W M, et al.2006. Physiological effects of exogenous nitric oxide in tomato seedlings under NaCl stress[J]. Journal of Chinese Agricultural Sciences, 15(3): 575-581.) [20] 许晓萱, 2019. NaCl胁迫对玉米幼苗生理及膜脂代谢影响的研究[D]. 硕士学位论文, 黑龙江八一农垦大学, 导师: 徐晶宇, pp. 17-20. (Xu X X, 2019. Effects of physiological and membrane lipid metabolism in maize seedlings under NaCl stress[D]. Thesis for M.S., Heilongjiang Bayi Agricultural University, Supervisor: Xu J Y, pp. 17-20 ). [21] 杨朴丽, 徐荣, 杨焱, 等. 2020. 低温胁迫对诺丽幼苗叶片光合荧光特性的影响[J]. 热带作物学报, 42(02): 455-464. (Yang P L, Xu R, Yang Y, et al.2020. Effects of low temperature stress on the photosynthetic fluorescence characteristics of Noni (Morindacitrifolia L.) seedling leaves[J]. Acta Tropical Crops, 42(02): 455-464.) [22] 张可炜, 王贤丽, 王雷, 等. 2007. 低温对不同磷水平下玉米叶片几种与光合作用有关的生理指标的影响[J]. 植物生理学报, 239(1): 93-97. (Zhang K W, Wang X L, Wang L, et al.2007. Effects of low temperature on some physiological indices of photosynthesis of maize (Zea mays L.) inbred lines under different phosphorus nutrition level[J]. Journal of Plant Physiology, 239(1): 93-97.) [23] 张文博. 2020. 低温胁迫下外源NO对黄瓜幼苗生长及多胺代谢的调控作用[D]. 硕士学位论文, 石河子大学, 导师: 崔金霞, pp. 15-17. (Zhang W B.2020. Effect of exogenous NO on the growth and polyamine metabolism of cucumber seedlings under low temperature stress[D]. Thesis for M.S., Shihezi University, Supervisor: Cui J X, pp. 15-17. ) [24] 赵剑颖, 宋晓莉, 杨蕊, 等. 2010. 4 ℃胁迫过程中大叶黄杨和北海道黄杨叶片抗寒生理生化指标的变化[J]. 北京农学院学报, 25(2): 57-61. (Zhao J Y, Song X L, Yang R, et al.2010. Variations of physiological and biochemical indexes of the leaf of Euonymus japonicus thunb. and Euonymus japonicus cv.'CuZhi' at 4 ℃ time course[J]. Journal of Beijing Agricultural College, 25(2): 57-61.) [25] 赵训超, 盖胜男, 魏玉磊, 等. 2020.低温胁迫下玉米根系生理变化及相关基因表达分析[J]. 农业生物技术学报, 28(1): 32-41. (Zhao X C, Ge S N, Wei Y L, et al.2020. Analysis of root physiology and related gene expression in maize (Zea mays) under low temperature stress[J]. Journal of Agricultural Biotechnology, (28): 32-41.) [26] 周福平, 柳青山, 张一中, 等. 2018. 低温胁迫对高粱幼苗叶绿素荧光参数的影响[J]. 种子, 37(9): 36-40. (Zhou F P, Liu Q S, Zhang Y Z, et al.2018. Effects of low temperature stress on chlorophyll fluorescence parameters of sorghum seedling[J]. Journal of Seeds, 37(9): 36-40.) [27] 周福平, 王坚强, 范娜, 等. 2020. 低温胁迫下高粱幼苗生理变化及耐冷综合评价[J]. 山西农业大学学报(自然科学版), 40(3): 45-53. (Zhou F P, Wang J Q, Fan N, et al.2020. Cold stress tolerance and physiological changes of sorghum seedling[J]. Journal of Shanxi Agricultural University (Natural Science Edition), 40(3): 45-53.) [28] 周亚峰. 2017. 甜瓜幼苗耐冷性评价体系构建及耐冷性相关位点的挖掘[D], 硕士学位论文,河南农业大学, 导师: 胡建斌, pp.24-31. (Zhou Y F, 2017. Construction of a system for evaluation of chilling tolerance in melon seedling and exploration of the loci related to chilling tolerance[D]. Thesis for M.S., Henan Agriculture University, Supervisor: Hu J B, pp.24-31.) [29] Chen X Y, Song F B, Liu F L, et al.2014. Effect of different arbuscular mycorrhizal fungi on growth and physiology of maize at ambient and low temperature regimes[J]. Scientific World Journal, 14(3): 1-7. [30] Czechowski T, Stitt M, Altmann T, et al.2005. Genome-wide identification and testing of superior reference genes for transcript normalization in Arabidopsis[J]. Plant Physiology, 139(1): 5-17. [31] Earl H J, Tollenaar M.1997. Maize left absorptance of photosynthetically active radiation and its estimation using a chlorophyll meter[J]. Crop Science, (37): 436-440. [32] Feng X, Lai Z X, Lin Y L, et al.2015. Genome-wide identification and characterization of the superoxide dismutase gene family in Musa acuminata cv. Tianbaojiao (aaa Group)[J]. BMC Genomics, 16(1): 1-16. [33] Foyer C H, Noctor G.2005. Redox homeostasis and antioxidant signaling: A metabolic interface between stress perception and physiological responses[J]. Plant Cell, 17(7): 1866-1875. [34] Francisco F, López-Cobollo R, Catalá R, et al.2002. A novel cold-inducible gene from Arabidopsis, Rci3, encodes a peroxidase that constitutes a component for stress tolerance[J]. The Plant Journal, 32(1): 13-24. [35] Fridovich.1975. The biology of oxgen redical[J]. Science, 201: 875-880. [36] Fryer M J, Oxborough K, Martin B, et al.1995. Factors associated with depression of photosynthetic quantum efficiency in maize at low growth temperature[J]. Plant Physiology, 108(2): 761-767. [37] Hodges D M, Delong J M, Forney C F, et al.1999. Improving the thiobarbituric acid-reactive-substabces assay for estimating lipid peroxidation in plant tissues containing anthocyanin and other interfering compounds[J]. Planta, 207(6): 604-611. [38] Grechkin A.1998. Recent developments in biochemistry of the plant lipoxygenase pathway[J]. Progress in Lipid Research, 37(5): 317-345. [39] Grene A R, Neval E, Lenwood H S.2002. Role of superoxide dismutases (SODs) in controlling oxidative stress in plants[J]. Journal of Experimental Botany, 53(372): 1331-1341. [40] Hur J, Jung K H, Lee C H, et al.2004. Stress-inducible OsP5CS2 gene is essential for salt and cold tolerance in rice[J]. Plant Science, 167(3): 417-426. [41] Mckersie B D, Chen Y, Beus M, et al.1993. Superoxide dismutase enhances tolerance of freezing stress in transgenic Alfalfa (Medicago sativa)[J]. Plant Physiology, 103(4): 1155-1163. [42] Su M, Li X F, Ma X Y, et al.2011. Cloning two P5CS genes from bioenergy sorghum and their expression profiles under abiotic stresses and meja treatment[J]. Plant Science, 181(6): 652-659. [43] Wei C, Cui Q, Zhang X Q, et al.2016. Three P5CS genes including a novel one from Lilium regale play distinct roles in osmotic, drought and salt stress tolerance[J]. Journal of Plant Biology, 59(5): 456-466. [44] Zhang J H, Jiang M Q.2002. Water stress-induced abscisic acid accumulation triggers the increased generation of reactive oxygen species and up-regulates the activities of antioxidant enzymes in maize leaves[J]. Journal of Experimental Botany, 19(379): 2401-2410. [45] Zhuang W F, Wu X J, Yang M, et al.2013. Influence of low-temperature stress on photosynthetic traits in maize seedlings[J]. Journal of Northeast Agricultural University, 20(3): 1-5.
