橡胶树<i>HbDHAR2</i>基因克隆及表达分析

doi:10.7668/hbnxb.20191875

摘要/Abstract

摘要： 脱氢抗坏血酸还原酶（DHAR）通过调节抗坏血酸水平在植物生长发育和抗逆过程中发挥重要作用。为明确橡胶树DHAR基因的序列特征、表达特性以及潜在的生物学功能，采用RT-PCR技术，从橡胶树中克隆了DHAR基因HbDHAR2。该基因开放阅读框为639 bp，编码212个氨基酸。预测HbDHAR2蛋白等电点为5.69，分子量为23.82 ku，是一个亲水蛋白。HbDHAR2蛋白含有谷胱甘肽S-转移酶（GST） N-端结构域和GST C-端DHAR结构域，属于GST超家族DHAR亚类。序列比对表明，HbDHAR2与木薯MeDHAR2和蓖麻RcDHAR2具有很高的一致性。组织表达谱分析显示，HbDHAR2在根、树皮、胶乳、成熟叶、衰老叶、嫩梢、雌花和雄花中均有表达，其中表达量最高的组织为嫩梢，表达量最低的组织为根。乙烯利（ETH）、茉莉酸甲酯（MeJA）、水杨酸（SA）、脱落酸（ABA）、过氧化氢（H₂O₂）、甲基紫精（MV）、低温、干旱和高盐等处理均能诱导HbDHAR2的表达。由此推测，HbDHAR2可能在橡胶树抵御非生物胁迫过程中发挥作用。结果为进一步研究橡胶树HbDHAR2基因的功能奠定了基础。

关键词: 橡胶树, 脱氢抗坏血酸还原酶, 克隆, 表达分析, 非生物胁迫

Abstract: Dehydroascorbate reductase (DHAR) plays an important role in plant growth, development, and abiotic stress tolerance by regulating ascorbic acid level. To reveal the sequence characteristic, expression pattern and possible biological function of DHAR gene in Hevea brasiliensis, HbDHAR2 was cloned from Hevea brasiliensis by RT-PCR. HbDHAR2 contained an open reading frame of 639 bp encoding a protein of 212 amino acid residues with an isoelectric point of 5.69 and a calculated molecular weight of 23.82 ku. HbDHAR2 was a hydrophilic protein. It contained a GST N-terminal domain and a GST C-terminal DHAR domain and belonged to the DHAR subclass of GST super family. Sequence alignment analysis indicated that HbDHAR2 showed high sequence identity with cassava MeDHAR2 and castor bean RcDHAR2. Tissue expression profiling revealed that HbDHAR2 was expressed in root, bark, latex, mature leaf, senescent leaf, new shoot, female flower and male flower of Hevea brasiliensis, with the highest expression in new shoot and the lowest expression in root. HbDHAR2 expression was induced by ethephon (ETH), methyl jasmonate (MeJA), salicylic acid (SA), abscisic acid (ABA), hydrogen peroxide (H₂O₂), methyl viologen (MV), cold, drought and salt treatments, indicating that HbDHAR2 might function in rubber tree tolerance to abiotic stress. These results lay a foundation for further studying the function of HbDHAR2 in Hevea brasiliensis.

Key words: Hevea brasiliensis, Dehydroascorbate reductase, Cloning, Expression analysis, Abiotic stress

中图分类号:

S794.1
Q78

李双江, 冯成天, 胡义钰, 袁坤, 刘进平, 王真辉, 刘辉. 橡胶树HbDHAR2基因克隆及表达分析[J]. 华北农学报, 2021, 36(3): 25-32. doi: 10.7668/hbnxb.20191875.

LI Shuangjiang, FENG Chengtian, HU Yiyu, YUAN Kun, LIU Jinping, WANG Zhenhui, LIU Hui. Cloning and Expression Analysis of HbDHAR2 Gene from Hevea brasiliensis[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2021, 36(3): 25-32. doi: 10.7668/hbnxb.20191875.

http://www.hbnxb.net/CN/Y2021/V36/I3/25

参考文献

[1] 张雪,陶磊,乔晟,杜秉昊,郭长虹. 谷胱甘肽转移酶在植物抵抗非生物胁迫方面的角色[J]. 中国生物工程杂志,2017,37(3):92-98.doi:10.13523/j.cb.20170313.
Zhang X,Tao L,Qiao S,Du B H,Guo C H. Roles of glutathione S-transferase in plant tolerance to abiotic stresses[J]. China Biotechnology,2017,37(3):92-98.
[2] Edwards R,Dixon D P,Walbot V. Plant glutathione S-transferases:enzymes with multiple functions in sickness and in health[J]. Trends in Plant Science,2000,5(5):193-198.doi:10.1016/s1360-1385(00)01601-0.
[3] Kumar S,Trivedi P K. Glutathione S-transferases:Role in combating abiotic stresses including arsenic detoxification in plants[J]. Frontiers in Plant Science,2018,9:751.doi:10.3389/fpls.2018.00751.
[4] Labrou N E,Papageorgiou A C,Pavli O,Flemetakis E. Plant GSTome:Structure and functional role in xenome network and plant stress response[J]. Current Opinion in Biotechnology,2015,32:186-194.doi:10.1016/j.copbio.2014.12.024.
[5] Lallement P A,Brouwer B,Keech O,Hecker A,Rouhier N. The still mysterious roles of cysteine-containing glutathione transferases in plants[J]. Frontiers in Pharmacology,2014,5:192.doi:10.3389/fphar.2014.00192.
[6] 王俊生,刘红占,范小芳,殷贵鸿,韩玉林,周琳,赵锦慧. 3种小麦DHAR蛋白高级结构建模与功能分析[J]. 分子植物育种,2016,14(9):2472-2480.doi:10.13271/j.mpb.014.002472.
Wang J S,Liu H Z,Fan X F,Yin G H,Han Y L,Zhou L,Zhao J H. Molecular structure and function aanalysis of three DHAR proteins from wheat (Triticum aestivum L.)[J]. Molecular Plant Breeding,2016,14(9):2472-2480.
[7] 成子硕,兰婷,李迪,杨海灵,曾庆银. 江南卷柏脱氢抗坏血酸还原酶的分子特性[J]. 生物工程学报,2011,27(1):76-84.doi:10.13345/j.cjb.2011.01.004.
Cheng Z S,Lan T,Li D,Yang H L,Zeng Q Y. Molecular characterizations of two dehydroascorbate reductases from Selaginella moellendorffii[J]. Chinese Journal of Biotechnology,2011,27(1):76-84.
[8] Liu Y J,Han X M,Ren L L,Yang H L,Zeng Q Y. Functional divergence of the glutathione S-transferase supergene family in Physcomitrella patens reveals complex patterns of large gene family evolution in land plants[J]. Plant Physiology,2013,161(2):773-786.doi:10.1104/pp.112.205815.
[9] 侯艳霞,张文鑫,杨婧,李小艳,郭艳. 植物脱氢抗坏血酸还原酶(DHAR)研究进展[J]. 农业技术与装备,2017(2):28-29,32.doi:10.3969/j.issn.1673-887X.2017.02.009.
Hou Y X,Zhang W X,Yang J,Li X Y,Guo Y. A review of progress on dehydroascorbate reductase(DHAR) in plants[J]. Agricultural Technology & Equipment,2017(2):28-29,32.
[10] Ding H Y,Wang B P,Han Y,Li S C. The pivotal function of dehydroascorbate reductase in glutathione homeostasis in plants[J]. Journal of Experimental Botany,2020,71(12):3405-3416.doi:10.1093/jxb/eraa107.
[11] Yamaguchi M,Joslyn M A. Purification and properties of dehydroascorbic acid reductase of peas (Pisum sativum)[J]. Archives of Biochemistry and Biophysics,1952,38(1):451-465.doi:10.1016/0003-9861(52)90051-9.
[12] Rahantaniaina M S,Li S C,Chatel-Innocenti G,Tuzet A,Issakidis-Bourguet E,Mhamdi A,Noctor G. Cytosolic and chloroplastic DHARs cooperate in oxidative stress-driven activation of the salicylic acid pathway[J]. Plant Physiology,2017,174(2):956-971.doi:10.1104/pp.17.00317.
[13] Urano J,Nakagawa T,Maki Y,Masumura T,Tanaka K,Murata N,Ushimaru T. Molecular cloning and characterization of a rice dehydroascorbate reductase[J]. FEBS Letters,2000,466(1):107-111.doi:10.1016/s0014-5793(99)01768-8.
[14] 袁进成,马海莲,宋晋辉,宋小青,刘颖慧. 玉米脱氢抗坏血酸还原酶基因的克隆和特性研究[J]. 西北植物学报,2013,33(9):1745-1749.
Yuan J C,Ma H L,Song J H,Song X Q,Liu Y H. Identification and characterization of a dehydroascorbate reductase gene in maize (Zea mays L.)[J]. Acta Botanica Boreali-Occidentalia Sinica,2013,33(9):1745-1749.
[15] 刘婧愉,滕瑞敏,李辉,刘昊,庄静. 茶树DHAR酶基因的克隆与非生物胁迫响应分析[J]. 园艺学报,2020,47(5):983-994.doi:10.16420/j.issn.0513-353x.2019-0684.
Liu J Y,Teng R M,Li H,Liu H,Zhuang J. Cloning and expression analysis under abiotic stress of the DHAR gene in Camellia sinensis[J]. Acta Horticulturae Sinica,2020,47(5):983-994.
[16] Chen Z,Gallie D R. The ascorbic acid redox state controls guard cell signaling and stomatal movement[J]. The Plant Cell,2004,16(5):1143-1162.doi:10.1105/tpc.021584.
[17] Chen Z,Gallie D R. Dehydroascorbate reductase affects leaf growth,development,and function[J]. Plant Physiology,2006,142(2):775-787.doi:10.1104/pp.106.085506.
[18] Terai Y,Ueno H,Ogawa T,Sawa Y,Miyagi A,Kawai-Yamada M,Ishikawa T,Maruta T. Dehydroascorbate reductases and glutathione set a threshold for high-light-induced ascorbate accumulation[J]. Plant Physiology,2020,183(1):112-122.doi:10.1104/pp.19.01556.
[19] Chen Z,Gallie D R. Dehydroascorbate reductase affects non-photochemical quenching and photosynthetic performance[J].The Journal of Biological Chemistry,2008,283(31):21347-21361.doi:10.1074/jbc.M802601200.
[20] Hasanuzzaman M,Bhuyan M H M B,Anee T I,Parvin K,Nahar K,Mahmud J A,Fujita M. Regulation of ascorbate-glutathione pathway in mitigating oxidative damage in plants under abiotic stress[J]. Antioxidants,2019,8(9):384.doi:10.3390/antiox8090384.
[21] Ushimaru T,Nakagawa T,Fujioka Y,Daicho K,Naito M,Yamauchi Y,Nonaka H,Amako K,Yamawaki K,Murata N. Transgenic Arabidopsis plants expressing the rice dehydroascorbate reductase gene are resistant to salt stress[J]. Journal of Plant Physiology,2006,163(11):1179-1184.doi:10.1016/j.jplph.2005.10.002.
[22] Le Martret B,Poage M,Shiel K,Nugent G D,Dix P J. Tobacco chloroplast transformants expressing genes encoding dehydroascorbate reductase,glutathione reductase,and glutathione-S-transferase,exhibit altered anti-oxidant metabolism and improved abiotic stress tolerance[J]. Plant Biotechnology Journal,2011,9(6):661-673.doi:10.1111/j.1467-7652.2011.00611.x.
[23] Shin S Y,Kim M H,Kim Y H,Park H M,Yoon H S. Co-expression of monodehydroascorbate reductase and dehydroascorbate reductase from Brassica rapa effectively confers tolerance to freezing-induced oxidative stress[J]. Molecules and Cells,2013,36(4):304-315.doi:10.1007/s10059-013-0071-4.
[24] Eltayeb A E,Kawano N,Badawi G H,Kaminaka H,Sanekata T,Morishima I,Shibahara T,Inanaga S,Tanaka K. Enhanced tolerance to ozone and drought stresses in transgenic tobacco overexpressing dehydroascorbate reductase in cytosol[J]. Physiologia Plantarum,2006,127(1):57-65.doi:10.1111/j.1399-3054.2006.00624.x.
[25] 张珊. 小麦脱氢抗坏血酸还原酶DHAR 基因的克隆与功能分析[D]. 杨凌:西北农林科技大学,2019.
Zhang S. Cloning and functional analysis of dehydroascorbate reductase DHAR gene in wheat[D]. Yangling:Northwest A & F University,2019.
[26] 刘辉,邓治,李德军. 巴西橡胶树HbSAP1 基因克隆及表达分析[J]. 热带作物学报,2016,37(3):510-516.doi:10.3969/j.issn.1000-2561.2016.03.014.
Liu H,Deng Z,Li D J. Cloning and expression analysis of HbSAP1 gene in Hevea brasiliensis[J]. Chinese Journal of Tropical Crops,2016,37(3):510-516.
[27] 刘辉,韦永选,邓治,代龙军,杨洪,李德军. 巴西橡胶树WRKY转录因子基因HbWRKY41 的克隆与表达分析[J]. 华北农学报,2017,32(5):91-96.doi:10.7668/hbnxb.2017.05.014.
Liu H,Wei Y X,Deng Z,Dai L J,Yang H,Li D J. Cloning and expression analysis of a WRKY transcription factor gene HbWRKY41 in Hevea brasiliensis[J]. Acta Agriculturae Boreali-Sinica,2017,32(5):91-96.
[28] Fomenko D E,Gladyshev V N. CxxS:fold-independent redox motif revealed by genome-wide searches for thiol/disulfide oxidoreductase function[J]. Protein Science,2002,11(10):2285-2296.doi:10.1110/ps.0218302.
[29] Akram N A,Shafiq F,Ashraf M. Ascorbic acid-a potential oxidant scavenger and its role in plant development and abiotic stress tolerance[J]. Frontiers in Plant Science,2017,8:613.doi:10.3389/fpls.2017.00613.
[30] Horemans N,Potters G,De Wilde L,Caubergs R J.Dehydroascorbate uptake activity correlates with cell growth and cell division of tobacco bright yellow-2 cell cultures[J]. Plant Physiology,2003,133(1):361-367.doi:10.1104/pp.103.022673.
[31] Verma V,Ravindran P,Kumar P P. Plant hormone-mediated regulation of stress responses[J]. BMC Plant Biology,2016,16:86.doi:10.1186/s12870-016-0771-y.

选择文件类型/文献管理软件名称

选择包含的内容

橡胶树HbDHAR2基因克隆及表达分析

Cloning and Expression Analysis of HbDHAR2 Gene from Hevea brasiliensis

PDF

可视化

被引次数

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics

本文评价

[1]	金一锋, 高岩松, 王琦, 王萌萌, 赵迪, 熊毅, 陈阳. *草地早熟禾蛋白激酶SnRK2.4* 基因克隆及非生物胁迫响应分析**[J]. 华北农学报, 2022, 37(5): 9-15.
[2]	曲东, 燕飞, 刘欣瑞, 康雪, 曾海涛, 张羽. *猕猴桃AcWRKY70基因序列克隆及其对溃疡病病原菌和激素处理表达模式分析*[J]. 华北农学报, 2022, 37(5): 166-173.
[3]	张冬梅, 冯亚艳, 修志君, 杨春芳, 杜美娥, 李得宙, 张笑宇. *硅酸钠诱导的马铃薯抗黑痣病StWRKY11基因克隆及生物信息学分析*[J]. 华北农学报, 2022, 37(5): 194-203.
[4]	郑玉斯, 王培, 郭颖, 白丽蓉, 喻达辉, 赵森. *合浦珠母贝PfCLEC17A* 基因的克隆及表达分析**[J]. 华北农学报, 2022, 37(5): 230-238.
[5]	肖士奎, 李芳, 张文婷, 吕淑芳, 史国安, 吴疆, 范丙友. *芍药ACS基因的克隆及其蛋白质原核表达*[J]. 华北农学报, 2022, 37(5): 36-44.
[6]	侯瑞泽, 侯玉翔, 许小勇, 李璿, 李梅兰. 普通白菜CYP同源基因克隆与表达分析[J]. 华北农学报, 2022, 37(5): 45-51.
[7]	吉祥, 宋志成, 韦晓玲, 杨煜, 隋炯明, 郭宝太. 马铃薯卷叶病毒与S病毒重组双CP多克隆抗体的制备[J]. 华北农学报, 2022, 37(4): 212-219.
[8]	张晓寒, 陈彦良, 马欣, 张珊珊, 韦善君. *沟叶结缕草ZmCOR410基因抗逆功能鉴定*[J]. 华北农学报, 2022, 37(4): 53-61.
[9]	王亚男, 梁岩岩, 严文培, 张银萍, 李强, 吴秀菊. *北细辛AhOMT1基因的克隆及原核表达分析*[J]. 华北农学报, 2022, 37(4): 62-70.
[10]	赵长江, 都梦翔, 宋巨奇, 徐尚缘, 贺琳, 徐晶宇, 杨克军, 李佐同. 玉米NRL基因家族鉴定与逆境表达分析[J]. 华北农学报, 2022, 37(4): 1-10.
[11]	闫留延, 李剑峰, 张世文, 张博, 王永芳, 张小梅, 祖超凡, 王振山, 桑璐曼, 何占祥, 贾小平, 董志平. *谷子SiPRR73基因的光温调控模式及非生物胁迫响应特性*[J]. 华北农学报, 2022, 37(4): 11-19.
[12]	程春红, 游经番, 蔡兆明, 叶春宏, 陈丽. *茎瘤芥BjuEAR1-1的克隆与表达分析*[J]. 华北农学报, 2022, 37(3): 37-43.
[13]	唐忠丽, 逯晓楠, 赵蕊, 刘庆华, 李梅兰, 雷逢进, 许小勇. *黄皮西葫芦类胡萝卜素合成酶基因的鉴定及PSY1和LCYE2克隆分析*[J]. 华北农学报, 2022, 37(3): 60-67.
[14]	张晋玉, 徐新娟, 晁毛妮, 张晓红, 吴向远, 高际涛, 黄中文. *大豆GmZAT12基因的克隆和表达分析*[J]. 华北农学报, 2022, 37(3): 1-7.
[15]	贾志强, 许云玉, 高雪, 陶宏征, 陈增敏, 刘雅婷, 李永忠. *辣椒CaWRKY30转录因子的克隆、亚细胞定位及番茄斑萎病毒侵染下的表达分析*[J]. 华北农学报, 2022, 37(3): 186-192.

模态框（Modal）标题

选择文件类型/文献管理软件名称

选择包含的内容

橡胶树HbDHAR2基因克隆及表达分析

Cloning and Expression Analysis of HbDHAR2 Gene from Hevea brasiliensis

PDF

可视化

被引次数

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics

本文评价