Abstract：Raffinose plays an important role in plant response to abiotic stress, and galactinol synthase (Gols) is an important enzyme involved in the synthesis of raffinose. In this study, Gols gene families were identified and analyzed in the whole genome, 8, 7, 14 and 14 Gols genes were identified in Gossypium arboreum, G. raimondi, G. hirsutum and G. barbadense, respectively. Evolutionary analysis showed that the Gols gene family could be divided into 7 subgroups. The resules of conservative motifs and gene structure analysis showed that gene structure and composition of conserved motifs were also similar between closely related family members. A lot of cis-elements relevant to plant response to stresses were distributed on promoter regions of GhGols. Tissue expression pattern analysis showed that some GhGols genes were predominantly expressed in specific tissues. Expression analysis showed that the expression of some GhGols was induced by PEG treatment, among which the expression of GhGols6 was significantly up-regulated under PEG treatment. The results of qPCR analysis showed that the transcription level of GhGols6 was also significantly induced under drought stress. Further the GhGols6 gene was silenced using virus induced gene silencing (VIGS) technology, the result showed that silencing GhGols6 decreased the tolerance of cotton to drought stress, indicating that GhGols6 was a positive regulator of cotton drought response. This study provides a reference for further study on the function of Gols gene in cotton drought response.
[1] 安汶, 铠常丹, 杨艺, 等. 2016. 利用 VIGS 技术沉默 GhBES1基因对棉花幼苗生理指标的影响[J]. 分子植物育种,14(3): 598-603. (An W K, Chang D, Yang Y, et al. 2016. The effects of silenced GhBES1 gene on physiological parameters of cotton seedlings by VIGS technique[J]. Molecular Plant Breeding, 14(3): 598-603. ) [2] 从青, 程龙军, 杨宁, 等. 2019. 植物肌醇半乳糖苷合酶的生理功能和调控机制[J]. 中国生物化学与分子生物学报, 35(11): 1193-1200. (Cong Q, Cheng L J, Yang N, et al. 2019. Physiological function and regulation mecha-nism of galactinol synthase in plants[J]. Chinese Journal of Biochemistry and Molecular Biology, 35(11): 1193-1200. ) [3] 柯裕州, 周金星, 卢楠, 等. 2009. 盐胁迫对桑树幼苗光合生理及叶绿素荧光特性的影响[J]. 林业科学研究, 22(2): 200-206. (Ke Y Z, Zhou J X, Lu N, et al. 2009. Effects of salinity on physiology and chlorophyll fluorescence characteristics of mullberry (Morus alba) seedlings[J]. Forestry Scientific Research, 22(2):200-206. ) [4] 兰宏兵, 余述燕, 朱庆莉, 等. 2021. 棉籽中棉子糖和棉酚综合利用研究进展[J]. 安徽化工, 47(03): 1-3. (Lan H B, Yu M L, Cao Y F, et al. 2021. Study progress on compre-hensive utilization of raffinose and gossypol in cotton-seed[J]. Anhui Chemical Industry, 47(03): 1-3. ) [5] 李腾宇, 汤孟玲, 曹跃芬, 等. 2019. 棉花抗旱研究进展[J]. 江苏农业科学, 47(20): 64-69. (Li T Y, Tang M L, Cao Y F, et al. 2019. Research progress on drought resistance of cotton[J]. Jiangsu Agricultural Sciences, 47(20): 64-69. ) [6] 刘爱丽, 魏梦园, 黎冬华, 等. 2020. 芝麻肌醇半乳糖苷合成酶基因 SiGolS6 的克隆及功能分析[J]. 中国农业科学, 53(17): 3432-3442. (Liu A L, Wei M Y, Li D H, et al. 2020. Cloning and function analysis of sesame galac-tional synthase gene SiGolS6 in Arabidopsis[J]. Chinese Agricultural Science, 53(17): 3432-3442. ) [7] 刘瑞达, 葛常伟, 王敏轩, 等. 2022. 陆地棉转录因子基因 GhMYB108 的克隆及其在抗旱中的作用[J]. 中国农业科学, 55(10): 1877-1890. (Liu R D, Ge C W, Wang M X, et al. 2022. Cloning and drought resistance analysis of transcription factor GhMYB108 in Gossypium hirsutum[J]. Chinese Agricultural Sciences, 55(10): 1877-1890. ) [8] 卢基来, 王志勇, 龙翔宇, 等. 2020. 巴西橡胶树 GolS 和 RS 家族基因理化特性与表达特征分析[J]. 分子植物育种, 18(20): 6665-6670. (Lu J L, Wang Z Y, Long X Y, et al. 2020. Characterization and expression profiles of ga-lactionl synthase and raffinose synthase in rubber tree[J]. Molecular Plant Breeding, 18(20): 6665-6670. ) [9] 邱爽, 张军, 何佳琦, 等. 2021. 大豆肌醇半乳糖苷合成酶基因 GmGolS 克隆及非生物胁迫表达分析[J]. 西南农业学报, 34(05): 945-949. (Qiu S, Zhang J, He J Q, et al. 2021. Cloning and expression of galaction synthase gene GmGolS in soybean under abiotic stress[J]. Jour-nal of Southwest Agriculture, 34(05): 945-949) [10] 吴灏. 2018. 旱涝胁迫对棉花生长和产量的影响及模拟[D]. 博士学位论文, 武汉大学, 导师: 王修贵, pp. 1-8. (Wu H. 2018. Simulation for the growth and yield of cotton in response to drought and water logging[D]. Thesis for Ph. D., WuhanUniversity, Suppervisor: Wang X G, pp. pp. 1-8. ) [11] 闫成仕. 2002. 水分胁迫下植物叶片抗氧化系统的响应研究进展[J]. 烟台师范学院学报(自然科学版), 18(3): 220-225. (Yan C S. 2002. Advances responses to water stress in plant leaves[J]. Yantai Normal University Journal (Natural Science Edition), 18(3): 220-225. ) [12] 杨霞, 李毅博, 白月梅, 等. 2016. 干旱条件下叶片非顺序衰老小麦顶二叶叶绿素荧光特性[J]. 干旱地区农业研究, 34(1): 173-179. (Yang X, Li Y B, Bai Y M, et al. 2016. Chlorophyll fluorescence characteristics of top two leaves in non-sequencial senescence wheat under drought condition[J]. Agricultural Research in Arid Ar-eas, 34(1): 173-179. ) [13] 易萌萌. 2018. 新疆沙冬青 AnGolS2 基因的克隆及表达调控研究[D]. 硕士学位论文, 沈阳农业大学, 导师:陈丽静, pp. 10-15. (Yi M M. 2018. Molecular cloming and ex-pression regulatory of AnGolS2 gene from Ammopiptan-thus mongolicus in Xinjiang[D]. Thesis for M. S., Shenyang Agricultural University, Suppervisor: Chen L J, pp. 10-15. ) [14] 张强, 杨玉珍, 彭方仁. 2009. 干旱胁迫下不同种源香椿可溶性蛋白的动态变化[J]. 安徽农业科学, 37(1): 65-66, 71. (Zhang Q, Yang Y Z, Peng F R, et al. 2009. Dynamic changes of soluble protein in different provenances of Toona sinensis under drought stress[J]. Journal of Anhui Agricultural Sciences, 37(1): 65-66, 71. ) [15] 赵小萌. 2020. 棉子糖寡糖提高番茄幼苗高温抗性的作用研究[D]. 博士学位论文, 沈阳农业大学, 导师: 齐明芳, pp. 7-9. (Zhang X M. 2020. Study on the efffect of raffi-nose oligosaccharides on high temperature resistance of tomato seedlings[D]. Thesis for Ph. D., Shenyang Agri-cultural University, Suppervisor: Qi M F, pp. 7-9. ) [16] 周建, 杨立峰, 郝峰鸽, 等. 2009. 低温胁迫对广玉兰幼苗光合及叶绿素荧光特性的影响[J]. 西北植物学报, 29(1): 136-142. (Zhou J, Yang L F, Hao F G, et al. 2009. Phto-synthesis and chlorophylly-fluorescence of Magnolia grandiflora seedlings under low temperature stress[J]. Northwest Botanical Journal, 22(2): 200-206. ) [17] Chen C, Chen H, Zhang Y, et al. 2020. Tbtools: an integrative toolkit developed for interactive analyses of big biologi-cal data[J]. Molecular Plant, 13(8): 1194-1202. [18] Dai Y, Shao M, Hannaway D, et al. 2009. Effect of Thrips tabaci on anatomical features, photosynthetic character-istics and chloro-phyll fluorescence of Hypericum samp-sonii leaves[J]. Crop Protect, 28(4): 327-332. [19] Egert A, Keller F, Peters S, et al. 2013. Abiotic stress-induced accumulation of raffinose in Arabidopsis leaves is medi-ated by a single raffinose synthase (RS5, At5g40390)[J]. BMC Plant Biology, 13: 218. [20] Elsayed A I, Rafudeen M S, D Golldack, et al. 2014. Physio-logical aspects of raffinose family oligosaccharides in plants: Protection against abiotic stress[J]. Plant Biolo-gy, 16(1): 1-8. [21] Fan Y, Yu M, Liu M, et al. 2017. Genome-wide identification, evolutionary and expression analyses of the galactinol synthase gene family in rapeseed and tobacco[J]. Inter-national Journal of Molecular Sciences, 18(12): 2768. [22] Gao M, Xu B, Wang Y, et al. 2020. Quantifying individual and interactive effects of elevated temperature and drought stress on cotton yield and fibre quality[J]. Jour-nal of Agronomy and Crop Science, 207(3): 422-436. [23] Gu L, Jiang T, Zhang C, et al. 2019. Maize HSFA2 and HSBP2 antagonistically modulate raffinose biosynthesis and heat tolerance in Arabidopsis[J]. The Plant Journal: for Cell and Molecular Biology, 100(1): 128-142. [24] Gu L, Zhang Y, Zhang M, et al. 2016. ZmGOLS2, a target of transcription factor ZmDREB2A, offers similar protec-tion against abiotic stress as ZmDREB2A[J]. Plant Mo-lecular Biology, 90(1-2): 157-170. [25] Hua C L, Young B M, Geun O H, et al. 2019. Poaceae type ii galactinol synthase 2 from antarctic flowering plant des-champsia antarctica and rice improves cold and drought tolerance by accumulation of raffinose family oligosac-charides in transgenic rice plants[J]. Plant and Cell Phys-iology, 61(1): 88-104. [26] Hulse S, Gois E. 2020. Identification, evolutionary and ex-pression analysis of the galactinol synthase (gols) genes in Panicum virgatum L. and Panicum hallii: An in silico approach[J]. Plant Gene, 244: 100262. [27] Li F, Fan G, Wang K, et al. 2014. Genome sequence of the cultivated cotton Gossypium arboreum[J]. Nature Genet-ics, 46(6): 567-572. [28] Liu Y D, Zhang L, Meng S D, et al. 2019. Expression of ga-lactinol synthase from Ammopiptanthus nanus in tomato improves tolerance to cold stress[J]. Journal of Experi-mental Botany, 71(1): 435-449. [29] Locke E L, Stushnoff C, Teixeira D, et al. 2006. Raffinose family oligosaccharides in protection from osmotic stresses and a review of plant responses to chilling, freezing, drought, and salinity[J]. Floriculture Ornamen-tal, Plant Biotechnology, 123-131 [30] Ma S, Lv J, Li X, et al. 2021. Galactinol synthase gene 4 (Cs-gols4) increases cold and drought tolerance in Cucumis sativus L. by inducing rfo accumulation and ros scaveng-ing[J]. Environmental and Experimental Botany, 185: 104406. [31] Maxwell K, Johnson G N. 2000. Chlorophyll fluorescence: A practi-Chor cal guide[J]. Journal of Experimental Bota-ny, 51(345): 659-668. [32] Miyazawa S I, Nishiguchi M, Kogawara S, et al. 2017. Isola-tion of the drought-and salt-responsive galactinol syn-thase (gols) gene from black poplar leaves and analysis of the transformants overexpressing gols[J]. Bulletin of the Forestry, Forest Products Research Institute, 16(2): 77-86. [33] Naguib W B, Divte P R, Chandra A, et al. 2021. Raffinose ac-cumulation and preferential allocation of carbon (14C) to developing leaves impart salinity tolerance in sugar beet[J]. Physiologia Plantarum, 173(4): 1421-1433. [34] van Rensburg H C J. 2016. The Arabidopsis GolS1 promotor as a potential biosensor for heat stress and fungal infection? [D]. Thesis for M. S., Stellenbosch University, Supervi-sor: Shaun W P, Bianke L, pp. 1-77. [35] Schneider T, Keller F. 2009. Raffinose in chloroplasts is syn-thesized in the cytosol and transported across the chloro-plast envelope[J]. Plant and Cell Physiology, 50(12): 2174-2182. [36] Shimosaka E, Ozawa K. 2015. Overexpression of cold-induc-ible wheat galactinol synthase confers tolerance to chill-ing stress in transgenic rice[J]. Breeding Science, 65(5): 363-371. [37] Shin Y K, Bhandari S R, Cho M C, et al. 2020. Evaluation of chlorophyll fluorescence parameters and proline content in tomato seedlings grown under different salt stress condition[J]. Horticulture, Environment and Biotechnol-ogy, 61(10): 433-443. [38] Sonali S, Sritama M, Papri B, et al. 2015. Significance of ga-lactinol and raffinose family oligosaccharide synthesis in plants[J]. Frontiers in Plant Science, 6(656): 656. [39] Song C, Chung W S, Lim C O, et al. 2016. Overexpression of heat shock factor gene hsfa3 increases galactinol levels and oxidative stress tolerance in Arabidopsis[J]. Mole-cules, Cells, 39(6): 477-483. [40] Vinson C C, Mota A, Porto B N, et al. 2020. Characterization of raffinose metabolism genes uncovers a wild arachis galactinol synthase conferring tolerance to abiotic stress-es[J]. Scientific Reports, 10(1): 15258. [41] Wang K, Wang Z, Li F, et al. 2012. The draft genome of a dip-loid cotton Gossypium raimondii[J]. Nature Genetics, 44(10): 1098-103. [42] Wang M, Tu L, Yuan D, et al. 2019. Reference genome se-quences of two cultivated allotetraploid cottons, Gossypi-um hirsutum and Gossypium barbadense[J]. Nature Genet-ics, 51(2): 224-229. [43] Zhang J X, Kirkham M B. 1996. Enzymatic responses of the ascorbate-glutathione cycle to drought in sorghum and sunflower plants[J]. Plant Science, 113(2): 139-147.
