紫花苜蓿bZIP转录因子MsbZIP1的克隆、表达特性和遗传转化分析

doi:10.11733/j.issn.1007-0435.2020.03.004

草地学报 ›› 2020, Vol. 28 ›› Issue (3): 613-622.DOI: 10.11733/j.issn.1007-0435.2020.03.004

紫花苜蓿bZIP转录因子MsbZIP1的克隆、表达特性和遗传转化分析

曹晓宇^1,2, 王学敏², 郭继承³, 崔苗苗², 张锦锦², 刘佼佼², 贺俊英¹

1. 内蒙古师范大学生命科学与技术学院, 内蒙古呼和浩特 010000;
2. 中国农业科学院北京畜牧兽医研究所, 北京 100193;
3. 北京绿京华园林工程有限公司, 北京 102209

收稿日期:2019-11-20 修回日期:2020-01-04 出版日期:2020-06-15 发布日期:2020-05-30
通讯作者: 贺俊英, 王学敏
作者简介:曹晓宇(1994-),女,内蒙古呼和浩特人,硕士研究生,主要从事紫花苜蓿抗逆性生理研究,E-mail:1498720999@qq.com;
基金资助:
国家自然科学基金面上项目（31872410）；现代农业产业技术体系牧草产业体系岗位科学家（CARS-34）；国家科技资源共享服务平台作物种质资源库（NICGR-78）共同资助

Cloning,Expression Characteristics and Genetic Transformation Analysis of A bZIP Transcription Factor MsbZIP1 in Alfalfa

CAO Xiao-yu^1,2, WANG Xue-min², GUO Ji-cheng³, CUI Miao-miao², ZHANG Jin-jin², LIU Jiao-jiao², HE Jun-ying¹

1. College of Life Science and Technology, Inner Mongolia Normal University, Hohhot, Inner Mongolia Autonomous Region 010000, China;
2. Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193, China;
3. Beijing Green Jinghua Garden Engineering Company Ltd., Beijing 102209, China

Received:2019-11-20 Revised:2020-01-04 Online:2020-06-15 Published:2020-05-30

摘要/Abstract

摘要： bZIP（Basic leucine zipper）是植物中一类非常重要的转录因子，参与植物从生长发育到抗性调控的多个生物学过程。为了了解紫花苜蓿（Medicago sativa）中bZIP转录因子的特性，解析紫花苜蓿MsbZIP1基因的生物学功能，从而阐述MsbZIP1基因响应紫花苜蓿抗逆调控机制，本研究利用RACE技术从紫花苜蓿中获得1个MsbZIP1基因的全长cDNA，该序列全长1 176 bp，编码361个氨基酸，预测分子量为42.3 kD，等电点为6.5。分析发现，该蛋白含有bZIP家族典型的BRLZ碱性结构域和亮氨酸拉链，属于bZIP家族蛋白。进化树分析表明，该蛋白属于bZIP转录因子C亚族，与拟南芥（Arabidopsis thaliana）的AtbZIP63具有很高的同源性，推测可能具有与该类蛋白相似的功能。qRT-PCR分析表明，MsbZIP1基因对干旱、高盐、高温、低温，以及脱落酸（Abscisic acid，ABA）和生长素（Auxin，IAA）处理都有不同程度的响应，推测该基因可能参与调控紫花苜蓿多种非生物胁迫。本试验通过构建表达载体PCAMBIA3301-MsbZIP1，以农杆菌介导的花序浸染法转化拟南芥，经后代筛选、扩繁和分子检测，得到7株超表达的转基因拟南芥。本研究第一次分离了紫花苜蓿C亚族bZIP转录因子，初步确定紫花苜蓿MsbZIP1基因响应多种逆境胁迫的反应，并获得了阳性转基因材料，并为进一步探索该类转录因子在紫花苜蓿抗逆性调控中的作用奠定了基础。

关键词: 紫花苜蓿, bZIP转录因子, 克隆, 表达, 转化

Abstract: The bZIP (Basic leucine zipper) genes is a type of very important transcription factors in plants. It is involved in many biological processes from growth and development to stress resistance regulation in plants. In order to understand the characteristics of bZIP transcription factors in alfalfa (Medicago sativa),to analyze the biological function of alfalfa MsbZIP1 gene,and to explain the mechanism of MsbZIP1 gene response to alfalfa stress resistance,the full-length cDNA of a MsbZIP1 gene was isolated from alfalfa in this research. Sequence analysis showed that the full-length of this gene is 1 176 bp,encoding a protein of 361 amino acids with a predicted molecular weight of 42.3 kD and an isoelectric point of 6.5. Smart analysis showed that MsbZIP1 contains a typical BRLZ motif (Basic region domain and leucine zipper domain) of the bZIP family. Phylogenetic tree analysis revealed that MsbZIP1 belongs to C subfamily of the bZIP family and is related closely to Aribidopsis bZIP63. qRT-PCR analysis indicated that MsbZIP1 gene responded to various abiotic stresses (dehydration,salinity,high temperature,low temperature and two kinds of hormones:ABA and Auxin),indicating that it may played an important role in the adaptation to abiotic stress in alfalfa. In this test,an over expression vector pCAMBIA3301-MsbZIP1 was then constructed and transformed into Arabidopsis thaliana by Agrobacterium-mediated floral dip method. Transgenic Arabidopsis plants over expressing MsbZIP1 was obtained by screening and molecular identification. In conclusion,this study isolated the C subfamily bZIP TF in forage legume alfalfa for the first time,preliminarily determined the response of alfalfa MsbZIP1 gene to multiple abiotic stress and successfully obtained the transgenic plants. The results lay a foundation for further exploring the role of this type of transcription factor in the regulation of alfalfa's stress resistance.

Key words: Alfalfa, bZIP transcription factor, Clone, Expression, Genetic transformation

中图分类号:

S541.9

曹晓宇, 王学敏, 郭继承, 崔苗苗, 张锦锦, 刘佼佼, 贺俊英. 紫花苜蓿bZIP转录因子MsbZIP1的克隆、表达特性和遗传转化分析[J]. 草地学报, 2020, 28(3): 613-622.

CAO Xiao-yu, WANG Xue-min, GUO Ji-cheng, CUI Miao-miao, ZHANG Jin-jin, LIU Jiao-jiao, HE Jun-ying. Cloning,Expression Characteristics and Genetic Transformation Analysis of A bZIP Transcription Factor MsbZIP1 in Alfalfa[J]. Acta Agrestia Sinica, 2020, 28(3): 613-622.

0
/ / 推荐

导出引用管理器 EndNote|Ris|BibTeX

链接本文: https://manu40.magtech.com.cn/Jweb_cdxb/CN/10.11733/j.issn.1007-0435.2020.03.004

https://manu40.magtech.com.cn/Jweb_cdxb/CN/Y2020/V28/I3/613

参考文献

[1] Nijhawan A,Jain M,Tyagi A K,et al. Genomic survey and gene expression analysis of the basic leucine zipper transcription factor family in rice[J]. Plant Physiology,2008,146(2):330-350
[2] 勒正伟. 蓖麻bZIP家族鉴定及与油脂基因互作分析[D]. 北京:中国科学技术大学,2014:5
[3] Silveira A B,Gauer L,Tomaz J P,et al. The Arabidopsis AtbZIP9 protein fused to the VP16 transcriptional activation domain alters leaf and vascular development[J]. Plant Science,2007,172:1148-1156
[4] Takahashi H,Kawakatsu T,Wakasa Y. A rice transmembrane bZIP transcription factor,OsbZIP39,Regulates the endoplasmic reticulum stress response[J]. Plant and Cell Physiology,2012,53:144-153
[5] Thalor S K,Berberich T,Lee S S. Deregulation of Sucrose-controlled translation of a bZIP-type transcription factor results in Sucrose accumulation in leaves[J]. PLOS ONE,2012,7(3):e33111
[6] Huang X,Ouyang X H,Yang P Y,et al. Arabidopsis FHY3 and HY5 Positively mediate Induction of COP1 Transcription in Response to Photomorphogenic UV-B Light[J]. Plant Cell,2012,24(11):4590-4606
[7] Preeti A,Vinay Kumar B,Paramjit K,et al. Genome-wide Analysis of bZIP Transcription Factors in wheat and Functional Characterization of a TabZIP under abiotic stress[J]. Scientific Reports,2019,9:4608-4019
[8] 刘莉. DREBs,bZIP转录因子与植物抗旱性研究进展[J]. 浙江农业科学,2013(1):98-102
[9] Wu S Y,hu P H,Jia B W,et al. A Glycine so jagroup S2 bZIP transcription factor[J]. Tree Physiology,2018,18:234
[10] Mouttet R,Escobar-Gutiérrez A,Eaquibet M,et al. Ban-ning of methyl bromide for seed treament:Could Dithylenchus dipsaci again become a major threat to alfalfa production in Europe[J]. Pest Management Science,2014,70(7):1017-1022
[11] 魏杰. 紫花苜蓿及其栽培技术[J]. 畜牧与饲料科学,2009,30(3):159-160
[12] Goldman Myla D,Dwyer Lauren,Coleman Rachael,et al. Patient-specific factors modulate leukocyte response in dimethyl fumarate treated MS patients[J]. PLOS ONE,2020,15(2):e0228617
[13] 许红梅,张丽军,刘淳. 农杆菌蘸花法侵染拟南芥的研究[J]. 北方园艺,2010(14):143-146
[14] Jakoby,Weisshar M,Dro ge-Laser B,et al. bZIP transcription factors in Arabidopsis[J]. Trends in Plant Science,2002,7(3):106-111
[15] Kim S,Kang J Y,Cho D I,et al. ABF2,an ABRE-binding bZIP factor,is an essential component of glucose signaling and its overexpression affects multiple stress tolerance[J]. Plant Journal,2004,40:75-87
[16] HOSSAIN M A,CHO J I,HAN M,et al. The ABRE-binding bZIP transcription factor OsABF2 is a positive regulator of abiotic stress and ABA signaling in rice[J]. Journal of Plant Physiology,2010,167(17):1512-1520
[17] Liao Y,Zou H F,Wei W,et al. Soybean GmbZIP44,GmbZIP62 and GmbZIP78 genes function as negative regulator of ABA signaling and confer salt and freezing tolerance in transgenic Arabidopsis[J]. Planta,2008,228(2):225-240
[18] Weisshaar B,Armstrong G A,Block A,et al. Light-inducible and constitutively expressed DNA-binding proteins recognizing a plant promoter element with functional relevance in light responsiveness[J]. EMBO Journal,1991,10:1777-1786
[19] Matiolli,Cleverson Carlos,Tomaz,et al. The Arabidopsis bZIP Gene AtbZIP63 is a Sensitive Integrator of Transient Abscisic Acid and Glucose Signals[J]. Plant Physiology,2011,157(2):692-705
[20] Alonso,Rosario,Oñate-Sánchez,et al. A Pivotal Role of the Basic Leucine Zipper Transcription Factor bZIP53 in the Regulation of Arabidopsis Seed Maturation Gene Expression Based on Heterodimerization and Protein Complex Formation(W)[J]. Plant Cell,2009,21(6):1747-1761
[21] 慧洁,徐恒,邱文怡,等. bZIP转录因子在植物生长发育及非生物逆境响应的作[J]. 浙江农业学报,2019,31(7):1205-1214
[22] JI X Y,LIU G F,LIU Y J,et al. The bZIP protein fromTamarix hispida,ThbZIP1,is ACGT elements binding factor that enhances abiotic stress signaling in transgenic Arabidopsis[J]. BMC Plant Biology,2013,13(1):151-152
[23] Wang J C,Xu H,Zhu Y,et al. OsbZIP58,a basic leucine zipper transcription factor,regulates starch biosynthesis in rice endosperm[J]. Journal of Experimental Botany,2013,64(11):3453-3466
[24] Liu C T,Wu Y B,Wang X P. bZIP transcription factor OsbZIP52/RISBZ5:a potential negative regulator of cold and drought stress response in rice[J]. Planta,2012,235(6):1157-1169
[25] 马璇,胡小倩,张颖翌,等.植物非生物胁迫中的bZIP转录因子[J/OL]. http://kns.cnki.net/kcms/detail/46.1068.S.1026.004.html,2019-06-24/2020-01-30
[26] Kang J,Choi H,Kim S Y. Arabidopsis basic leucine zipper proteins that mediate stress responsive abscisic acid signaling[J]. Plant Cell,2002,14(2):343-357
[27] Bi C,Ma Y,Wu Z,et al. Arabidopsis ABI5 plays a role in regulating ROS homeostasis by activating CATALASE1 transcription in seed germination[J]. Plant Molecular Biology,2017,94(1-2):197-213
[28] Fujita Y,Fujita M,Satoh R,et al. AREB1 is a Transcription Activator of Novel ABRE-Dependent ABA Signaling that Enhances Drought Stress Tolerance in Arabidopsis[J]. Plant Cell,2005,17(12):3470-3488
[29] Ma H Z,Liu C,Li Z X. ZmbZIP4 Contributes to Stress Resistance in Maize by Regulating ABA Synthesis and Root Development[J]. Plant Physiology,2018,178:753-770

紫花苜蓿bZIP转录因子MsbZIP1的克隆、表达特性和遗传转化分析

Cloning,Expression Characteristics and Genetic Transformation Analysis of A bZIP Transcription Factor MsbZIP1 in Alfalfa

PDF

可视化

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]	王静, 刘晓静, 程甜甜, 童长春, 汪雪. 不同氮效率紫花苜蓿叶特征及其产量效应研究[J]. 草地学报, 2021, 29(9): 1941-1949.
[2]	李鑫, 高佩然, 边秀举, 王丽宏, 李会彬, 刘畅, 董康挺, 孙鑫博. 匍匐翦股颖叶肉细胞原生质体瞬时表达体系的建立[J]. 草地学报, 2021, 29(9): 2091-2097.
[3]	孟宣辰, 马鹏程, 马杰, 段珍, 韩阳阳, 张吉宇. 黄土高原半干旱区紫花苜蓿覆膜垄沟和施肥效应研究[J]. 草地学报, 2021, 29(9): 2098-2106.
[4]	朱海霞. 多孢木霉HZ-31菌株胁迫下野燕麦内参基因的选择[J]. 草地学报, 2021, 29(8): 1666-1674.
[5]	陈雷, 暴雪艳, 郭刚, 霍文婕, 李清宏, 许庆方, 王聪, 刘强. 单宁酸和乳酸菌对紫花苜蓿青贮品质和体外瘤胃发酵的影响[J]. 草地学报, 2021, 29(8): 1853-1858.
[6]	金一锋, 陈阳, 齐欣, 高岩松, 金忠民, 赵清峰, 王琦. 草地早熟禾NRT1/PTR FAMILY 8.3基因的克隆及表达分析[J]. 草地学报, 2021, 29(7): 1397-1405.
[7]	李春萍, 邱轶辉, 夏厚胤, 谭璐娜, 许环宇, 张瀚文, 高景慧. 紫花苜蓿多父本F₁代低蒸腾及产量性状的隶属函数分析[J]. 草地学报, 2021, 29(7): 1416-1422.
[8]	李天琦, 林志玲, 卢轩, 黄佳媛, 杨宁, 沃野, 高凯, 柳小妮, 王显国. 灌溉量对科尔沁沙地紫花苜蓿土壤速效养分分布及淋失的影响[J]. 草地学报, 2021, 29(7): 1454-1461.
[9]	李鹏珍, 赵得琴, 邓波, 杨军, 赵国华, 赵亮. 生物炭与干旱胁迫对接种紫花苜蓿光合效率及生长的影响[J]. 草地学报, 2021, 29(6): 1257-1264.
[10]	王文静, 麻冬梅, 赵丽娟, 马巧利. 2,4-表油菜素内酯对盐胁迫下紫花苜蓿生理指标及根系离子积累的影响[J]. 草地学报, 2021, 29(6): 1363-1368.
[11]	潘新怡, 史昆, 龚攀, 韦宝, 吴欣明, 王赞. 紫花苜蓿花青苷合成的转录组分析[J]. 草地学报, 2021, 29(5): 866-875.
[12]	其美拉姆, 普布卓玛, 崔彤彤, 张新飞, 姜惠娜, 付娟娟, 呼天明. 外源钙对低温胁迫下西藏野生垂穗披碱草生理及相关基因表达的影响[J]. 草地学报, 2021, 29(5): 919-928.
[13]	黄荣, 姚博, 张玉娟, 贾倩英, 李向阳, 张宏, 张振粉. 成团泛菌CQ10脂多糖对紫花苜蓿种子萌发及苗期生长的影响[J]. 草地学报, 2021, 29(5): 965-971.
[14]	朱瑞婷, 牛奎举, 张娣君, 杨阳, 宋云, 马晖玲. 外源2,4-表油菜素内酯对镉胁迫下草地早熟禾叶绿素代谢的影响[J]. 草地学报, 2021, 29(4): 635-643.
[15]	徐洪雨, 甄莉丽, 李钰莹, 董宽虎, 李向林. 低温干旱环境对紫花苜蓿根颈耐寒性的影响[J]. 草地学报, 2021, 29(4): 724-733.