欢迎访问作物学报,今天是

作物学报 ›› 2015, Vol. 41 ›› Issue (09): 1343-1352.doi: 10.3724/SP.J.1006.2015.01343

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

野生大豆碳酸盐胁迫应答基因GsMIPS2的克隆及功能分析

陈晨1, 孙晓丽2, 刘艾林1, 端木慧子1, 于洋1, 肖佳雷1, 朱延明1, *   

  1. 1 东北农业大学生命科学学院, 黑龙江哈尔滨 150030; 2 黑龙江八一农垦大学农学院, 黑龙江大庆 163319
  • 收稿日期:2015-01-20 出版日期:2015-09-12 网络出版日期:2015-09-12
  • 通讯作者: 朱延明, E-mail:ymzhu@neau.edu.cn, Tel: 18645035310
  • 作者简介:第一作者联系方式: E-mail:a09080003@163.com, Tel: 18045476676
  • 基金资助:
    本研究由国家转基因生物新品种培育重大专项(2011ZX08004-002), 国家自然科学基金项目(31171578), 黑龙江省高校科技创新团队建设计划项目(2011TD055)和国家基础科学人才培养基金项目(J1210069)资助

Cloning and Functional Analysis of Glycine soja Bicarbonate Stress Responsive Gene GsMIPS2

CHEN Chen1, SUN Xiao-Li2, LIU Ai-Lin1, DUAN-MU Hui-Zi1, YU Yang1, XIAO Jia-Lei1, ZHU Yan-Ming1, *   

  1. 1 College of Life Sciences, Northeast Agricultural University, Harbin 150030, China; 2 College of Agronomy, Heilongjiang Bayi Agricultural University, Daqing 163319, China
  • Received:2015-01-20 Published:2015-09-12 Published online:2015-09-12

摘要: 为挖掘野生大豆(Glycine soja L. G07256)耐碳酸盐关键功能基因, 利用前期高通量转录组测序数据, 从构建的碳酸盐胁迫基因表达谱中, 选取了一个碳酸盐胁迫下显著上调表达的肌醇-1-磷酸合酶类基因。采用同源克隆的方法, 获得该基因的全长cDNA, 命名为GsMIPS2。实时荧光定量PCR结果显示该基因受碳酸盐胁迫诱导表达, 并且其表达量具有组织特异性。将GsMIPS2基因转化拟南芥, 并结合拟南芥中T-DNA插入突变体atmips2来验证其耐碳酸盐功能。结果表明, 碳酸盐胁迫条件下, GsMIPS2超量表达拟南芥种子萌发率显著高于野生型, 而拟南芥突变体atmips2种子萌发率显著低于野生型。上述结果表明, GsMIPS2基因在植物应答碳酸盐胁迫过程中起重要作用。

关键词: 野生大豆, GsMIPS2基因, 肌醇-1-磷酸合成酶, 碳酸盐胁迫

Abstract: To identify the key functional genes in response to bicarbonate stress in Glycine soja L. G07256, we constructed a gene expression profile under bicarbonate treatment using high throughput RNA-seq data, from which we selected a myo-inositol- 1-phosphate synthase gene whose expression was significantly induced by bicarbonate stress. By homology-based cloning, we acquired its full length cDNA and termed as GsMIPS2. The results of quantitative real-time PCR demonstrated the bicarbonate stress induced expression of GsMIPS2 and its tissue expression specificity. We also verified the function of GsMIPS2 in bicarbonate responses by using the GsMIPS2 transgenic Arabidopsis combined with the T-DNA insertion line atmips2. We revealed that the germination rate of GsMIPS2 overexpression lines was significantly higher, while that of atmips2 mutant line was much lower than that of wild type under bicarbonate stress. These results indicate the positive role of GsMIPS2 in plant bicarbonate stress responses.

Key words: Glycine soja, GsMIPS2, Myo-inositol-1-phosphate synthase, Bicarbonate stress

[1] Kawanabe S, Zhu T C. Degeneration and conservation of Aneurolepisium chinense grassland in northern China. J Jpn Grassl Sci , 1991, 37: 91-99
[2] 李彬, 王志春, 孙志高, 陈渊, 杨福. 中国盐碱地资源与可持续利用研究. 干旱地区农业研究, 2005, 23(2): 154-158 Li B, Wang Z C, Sun Z G, Chen Y, Yang F. Resources and sustainable resource exploitation of salinized land in China. Agric Res Arid Areas , 2005, 23(2): 154-158 (in Chinese with English abstract)
[3] Nakashima K, Ito Y, Yamaguchi-Shinozaki K. Transcriptional regulatory networks in response to abiotic stresses in Arabidopsis and grasses. Plant Physiol , 2009, 149: 88-95
[4] 李向华, 王克晶, 李福山, 严茂粉. 野生大豆( Glycine soja )研究现状与建议. 大豆科学, 2005, 24: 305-309 Li X H, Wang K J, Li F S, Yan M F. Research progress of wild soybean ( Glycine soja ) and suggestions for improving its effective utilization and protection. Soybean Sci , 2005, 24: 305-309 (in Chinese with English abstract)
[5] Loewusa F A, Murthy P P N. Myo -inositol metabolism in plants. Plant Sci , 2000, 150: 1-19
[6] Nelson D E, Rammesmayer G, Bohnert H J. Regulation of cell-specific inositol metabolism and transport in plant salinity tolerance. Plant Cell , 1998, 10: 753-764
[7] Das-Chatterjee A, Goswami L, Maitra S, Dastidar K G, Ray S, Majumde A L. Introgression of a novel salt-tolerant L- myo - inositol 1-phosphate synthase from Porteresia coarctata (Roxb.) Tateoka ( PcINO1 ) confers salt tolerance to evolutionary diverse organisms. FEBS Lett , 2006, 580: 3980-3988
[8] Patra B, Ray S, Richter A, Majumder A L. Enhanced salt tolerance of transgenic tobacco plants by co-expression of PcINO1 and McIMT1 is accompanied by increased level of myo -inositol and methylated inositol. Protoplasma , 2010, 245: 143-152
[9] Kaur H, Verma P, Petla B P, Rao V, Saxena S C, Majee M. Ectopic expression of the ABA-inducible dehydration-responsive chickpea L- myo -inositol 1-phosphate synthase 2 ( CaMIPS2 ) in Arabidopsis enhances tolerance to salinity and dehydration stress. Planta , 2013, 237: 321-335
[10] Joshi R, Ramanarao M V, Baisakh N. Arabidopsis plants constitutively overexpressing a myo -inositol 1-phosphate synthase gene ( SaINO1 ) from the halophyte smooth cordgrass exhibits enhanced level of tolerance to salt stress. Plant Physiol Biochem , 2013, 65: 61-66
[11] Hegeman C E, Good L L, Grabau E A. Expression of D- myo -inositol-3-phosphate synthase in soybean. Implications for phytic acid biosynthesis. Plant Physiol , 2001, 125: 1941-1948
[12] Kido E A, Ferreira Neto J R, Silva R L, Belarmino L C, Bezerra Neto J P, Soares-Cavalcanti N M, Pandolfi V, Silva M D, Nepomuceno A L, Benko-Iseppon A M. Expression dynamics and genome distribution of osmoprotectants in soybean: identifying important components to face abiotic stress. BMC Bioinform , 2013, 14(suppl) 1: S7
[13] Willems E, Leyns L, Vandesompele J. Standardization of real-time PCR gene expression data from independent biological replicates. Anal Biochem , 2008, 379: 127-129
[14] Nour-Eldin H H, Hansen B G, Norholm M H, Jensen J K, Halkier B A. Advancing uracil-excision based cloning towards an ideal technique for cloning PCR fragments. Nucl Acids Res , 2006, 34: e122
[15] Clough S J, Bent A F. Floral dip: a simplified method for Agrobacterium -mediated transformation of Arabidopsis thaliana . Plant J , 1998, 16:735-743
[16] Lu B B, Du Z, Ding R X, Zhang L, Yu X J, Liu C H, Chen W S. Cloning and characterization of a differentially expressed phenylalanine ammonialyse gene ( liPAL ) after genome duplication from tetraploid Isatis indigotica fort. J Integr Plant Biol , 2006, 48: 1439-1449
[17] Cui M, Liang D, Wu S, Ma F W, Lei Y S. Isolation and developmental expression analysis of L- myo -inositol-1-phosphate synthase in four Actinidia species. Plant Physiol Biochem , 2013, 73: 351-358
[18] Rao P S, Mishra B, Gupta S R, Rathore A. Reproductive stage tolerance to salinity and alkalinity stresses in rice genotypes. Plant Breed , 2008, 127: 256-261
[19] Shi D C, Sheng Y M. Effect of various salt-alkaline mixed stress conditions on sunflower seedlings and analysis of their stress factors. Environ Exp Bot , 2005, 54: 8-21
[20] Shi D C, Yin L J. Difference between salt (NaCl) and alkaline (Na 2 CO 3 ) stresses on Puccinellia tenuiflora (Griseb.) Scribn et Merr. plants. Acta Bot Sin , 1993, 3: 144-149
[21] Majee M, Maitra S, Dastidar K G, Pattnaik S, Chatterjee A, Hait N C, Das K P, Majumder A L. A novel salt-tolerant L- myo - inositol-1-phosphate synthase from Porteresia coarctata (Roxb.) Tateoka, a halophytic wild rice: molecular cloning, bacterial overexpression, characterization, and functional introgression into tobacco-conferring salt tolerance phenotype. J Biol Chem , 2004, 279: 28539-28552
[22] Ghosh Dastidar K, Maitra S, Goswami L, Roy D, Das K P, Majumder A L. An insight into the molecular basis of salt tolerance of L- myo -inositol 1-P synthase ( PcINO1 ) from Porteresia coarctata (Roxb.) Tateoka, a halophytic wild rice. Plant Physiol , 2006, 140: 1279-1296
[23] Ge Y, Li Y, Zhu Y M, Bai X, Lv D K, Guo D, Ji W, Cai H. Global transcriptome profiling of wild soybean ( Glycine soja ) roots under NaHCO 3 treatment. BMC Plant Biol , 2010, 10: 153
[24] Johnson M D, Sussex I M. 1L- myo -inositol 1-phosphate synthase from Arabidopsis thaliana . Plant Physiol , 1995, 107: 613-619
[25] Wongkaew A, Nakasathien S, Srinives P. Isolation and characterization of D- myo -inositol-3-phosphate synthase from mungbean ( Vigna radiata ). Plant Mol Biol Rep , 2009, 28: 122-127
[26] Majumdera A L, Johnsonb M D, Henry S A. 1L- myo -inositol- 1-phosphate synthase. Biochim Biophys Acta , 1997, 1348: 245-256
[27] Zhu D, Li R, Liu X, Sun M, Wu J, Zhang N, Zhu Y. The positive regulatory roles of the TIFY10 proteins in plant responses to alkaline stress. PLoS One , 2014, 9: e111984
[28] Yoshida K T, Wada T, Koyama H, Mizobuchi-Fukuoka R, Naito S. Temporal and spatial patterns of accumulation of the transcript of myo -inositol-1-phosphate synthase and phytin-containing particles during seed development in rice. Plant Physiol , 1999, 119: 65-72
[29] Boominathan P, Shukla R, Kumar A, Manna D, Negi D, Verma P K, Chattopadhyay D. Long term transcript accumulation during the development of dehydration adaptation in Cicer arietinum . Plant Physiol , 2004, 135: 1608-1620
[30] Raboy V. Myo -inositol-1,2,3,4,5,6-hexakisphosphate. Phytochemistry , 2003, 64: 1033-1043
[31] Saxena S C, Salvi P, Kaur H, Verma P, Petla B P, Rao V, Kamble N, Majee M. Differentially expressed myo -inositol monophosphatase gene ( CaIMP ) in chickpea ( Cicer arietinum L.) encodes a lithium-sensitive phosphatase enzyme with broad substrate specificity and improves seed germination and seedling growth under abiotic stresses. J Exp Bot , 2013, 64: 5623-5639
[1] 陈影,张晟瑞,王岚,王连铮,李斌,孙君明. 野生和栽培大豆种质油脂组成特点及其与演化的关系[J]. 作物学报, 2019, 45(7): 1038-1049.
[2] 朱娉慧**,陈冉冉**,于洋,宋雪薇,李慧卿,杜建英,李强,丁晓东,朱延明*. 碱胁迫相关基因GsWRKY15的克隆及其转基因苜蓿的耐碱性分析[J]. 作物学报, 2017, 43(09): 1319-1327.
[3] 王吴彬,何庆元,杨红燕,向仕华,邢光南,赵团结,盖钧镒. 大豆结荚习性、荚色和种皮色相关野生片段分析[J]. 作物学报, 2013, 39(07): 1155-1163.
[4] 范虎,文自翔,王春娥,王芳,邢光南,赵团结,盖钧镒. 中国野生大豆群体农艺加工性状与SSR关联分析和特异材料的遗传构成[J]. 作物学报, 2013, 39(05): 775-788.
[5] 才华, 朱延明, 李勇, 柏锡, 纪巍, 王冬冬, 孙晓丽. 野生大豆转录因子GsNAC20基因的分离及胁迫耐性分析[J]. 作物学报, 2011, 37(08): 1351-1359.
[6] 肖鑫辉, 李向华, 刘洋, 张应, 王克晶. 高盐碱胁迫下野生大豆(Glycine soja)体内离子积累的差异[J]. 作物学报, 2011, 37(07): 1289-1300.
[7] 王希,李勇,朱延明,柏锡,才华,纪巍. 野生大豆胁迫应答膜联蛋白基因的克隆及胁迫耐性分析[J]. 作物学报, 2010, 36(10): 1666-1673.
[8] 樊金萍;柏锡;李勇;纪巍;王希;才华;朱延明. 野生大豆S-腺苷甲硫氨酸合成酶基因的克隆及功能分析[J]. 作物学报, 2008, 34(09): 1581-1587.
[9] 文自翔;赵团结;郑永战;刘顺湖;王春娥;王芳;盖钧镒. 中国栽培和野生大豆农艺及品质性状与SSR标记的关联分析 II. 优异等位变异的发掘[J]. 作物学报, 2008, 34(08): 1339-1349.
[10] 文自翔;赵团结;郑永战;刘顺湖;王春娥;王芳;盖钧镒. 中国栽培和野生大豆农艺品质性状与SSR标记的关联分析 I. 群体结构及关联标记[J]. 作物学报, 2008, 34(07): 1169-1178.
[11] 张跃强;关荣霞;刘章雄;常汝镇;姚源松;邱丽娟. 利用大豆核心种质部分样本鉴定28K和30K过敏蛋白缺失材料[J]. 作物学报, 2006, 32(03): 324-329.
[12] 李福山;李向华. 野生大豆在自然界中光温反应的规律[J]. 作物学报, 2003, 29(05): 670-675.
[13] 董英山;庄炳昌;赵丽梅;孙寰;张明;何孟元. 中国野生大豆遗传多样性中心[J]. 作物学报, 2000, 26(05): 521-527.
[14] 盖钧镒;许东河;高忠;岛本义也;阿部纯;福士泰史;北岛俊二. 中国栽培大豆和野生大豆不同生态类型群体间遗传演化关系的研究[J]. 作物学报, 2000, 26(05): 513-520.
[15] 杨光宇;郑惠玉;韩春凤. 栽培大豆(G.max)×半野生大豆(G.gracilis)后代主要农艺性状遗传参数的初步分析[J]. 作物学报, 1992, 18(06): 439-446.
Viewed
Full text


Abstract

Cited

  Shared   
  Discussed   
No Suggested Reading articles found!