生物技术通报 ›› 2021, Vol. 37 ›› Issue (2): 32-39.doi: 10.13560/j.cnki.biotech.bull.1985.2020-0744
收稿日期:
2020-06-17
出版日期:
2021-02-26
发布日期:
2021-02-26
作者简介:
陈莹,女,硕士,研究方向:牧草育种;E-mail: 基金资助:
CHEN Ying(), CHEN Xi, WANG Qian, WANG Xiao-li()
Received:
2020-06-17
Published:
2021-02-26
Online:
2021-02-26
摘要:
逆境胁迫蛋白(Universal stress protein,USP)参与干旱、高温、氮胁迫和高盐等多种逆境胁迫反应。为研究逆境胁迫蛋白USP在高羊茅中的表达模式,采用cDNA末端快速扩增技术从高羊茅叶片中克隆USP基因,命名为FaUSP。结果表明,FaUSP全长为844 bp,含501 bp的开放阅读框,共编码166个氨基酸,具有USP家族典型的USPA结构域。经系统进化树分析,高羊茅FaUSP蛋白与禾本科植物小麦、大麦、山羊草的USP蛋白亲缘关系较近。荧光定量PCR表明,高羊茅FaUSP受干旱、高温、氮胁迫及高盐胁迫的诱导表达上调,构建FaUSP超量表达载体,利用农杆菌侵染法转化菊苣,干旱胁迫下过量表达FaUSP菊苣可溶性蛋白、GR、SOD、POD、CAT的合成显著高于野生型,表明FaUSP的超量表达可以增强菊苣的抗旱能力,推测该基因与抗旱性相关。
陈莹, 陈锡, 王茜, 王小利. 高羊茅逆境胁迫蛋白基因FaUSP的克隆、表达及生物学功能分析[J]. 生物技术通报, 2021, 37(2): 32-39.
CHEN Ying, CHEN Xi, WANG Qian, WANG Xiao-li. Cloning,Expression and Biological Function Analysis of Universal Stress Protein in Festuca arundinacea[J]. Biotechnology Bulletin, 2021, 37(2): 32-39.
引物名称 | 引物序列(5'-3') | 扩增类别 |
---|---|---|
FU1 | CAATGGAAGGCTCAGA | 5' RACE |
FU2 | AGAGAGGGGGATCAAAGG | |
FU3 | GGATGGTCTCCCTCTGCT | |
FU4 | CATCCCCATGAGCTGCCTGGTTATC | 3' RACE |
FU5 | AAAGGGTGCTCTTAGGCAGCGTCAG | |
FU-fwd1 | AGAGGTGGGTTGGCCTGGC | 中间片段PCR扩增 |
FU-rev1 | TCGCTGGCTTGACAACTGTG | |
UBI-fwd1 | CACCTCGATCACCCACCTCT | UBI内参引物 |
UBI-rev1 | AGGGTCTCCGATAACCTCCA | |
FU-fwd2 | CCTTTGATCCCCCTCTCTGAA | 荧光定量PCR引物 |
FU-rev2 | AAACCACCGTGACCTCCTTCT |
表1 高羊茅FaUSP克隆及荧光定量PCR的引物序列
引物名称 | 引物序列(5'-3') | 扩增类别 |
---|---|---|
FU1 | CAATGGAAGGCTCAGA | 5' RACE |
FU2 | AGAGAGGGGGATCAAAGG | |
FU3 | GGATGGTCTCCCTCTGCT | |
FU4 | CATCCCCATGAGCTGCCTGGTTATC | 3' RACE |
FU5 | AAAGGGTGCTCTTAGGCAGCGTCAG | |
FU-fwd1 | AGAGGTGGGTTGGCCTGGC | 中间片段PCR扩增 |
FU-rev1 | TCGCTGGCTTGACAACTGTG | |
UBI-fwd1 | CACCTCGATCACCCACCTCT | UBI内参引物 |
UBI-rev1 | AGGGTCTCCGATAACCTCCA | |
FU-fwd2 | CCTTTGATCCCCCTCTCTGAA | 荧光定量PCR引物 |
FU-rev2 | AAACCACCGTGACCTCCTTCT |
[1] |
VanBogelen RA, Hutton ME, Neidhardt FC. Gene-protein database of Escherichia coli K-12:edition 3[J]. Electrophoresis, 1990,11(12):1131-1166.
doi: 10.1002/elps.1150111205 URL pmid: 1965305 |
[2] |
Freestone P, Nystrom T, Trinei M, et al. The universal stress protein, UspA, of Escherichia coli is phosphorylated in response to stasis[J]. Journal of Molecular Biology, 1997,274:318-324.
doi: 10.1006/jmbi.1997.1397 URL pmid: 9405142 |
[3] |
David K, Joshua B, Douglas W, et al. Arabidopsis proteins containing similarity to the universal stress protein domain of bacteria[J]. Plant Physiology, 2003,131(3):1209-1219.
doi: 10.1104/pp.102.016006 URL pmid: 12644671 |
[4] |
Yang ML, Che SY, Zhang YX, et al. Universal stress protein in Malus sieversii confers enhanced drought tolerance[J]. Journal of Plant Research, 2019,132:825-837.
doi: 10.1007/s10265-019-01133-7 URL pmid: 31482250 |
[5] | Maqbool A, Zahur M, Husnain T, et al. GUSP1 and GUSP2 two drought-responsive genes in Gossypium arboreturn have homology to universal stress protein[J]. Plant Mol Biol Rep, 2009,27:109-114. |
[6] |
Jung YJ, Melencion SMB, Lee ES, et al. Universal stress protein exhibits a redox-dependent chaperone function in Arabidopsis and enhances plant tolerance to heat shock and oxidative stress[J]. Frontiers in Plant Science, 2015,6:1141.
doi: 10.3389/fpls.2015.01141 URL pmid: 26734042 |
[7] |
Bhuria M, Goel P, Kumar S, et al. The promoter of AtUSP is co-regulated by phytohormones and abiotic stresses in Arabidopsis thaliana[J]. Frontiers in Plant Science, 2016,7:1957.
doi: 10.3389/fpls.2016.01957 URL pmid: 28083000 |
[8] | 黄姗, 王伟旗, 侯文胜. 大豆Usp1基因的克隆和表达分析[J]. 大豆科学, 2012,31(4):546-551. |
Huang S, Wang WQ, Hou WS. Cloning and expression analysis of usp1 gene from soybean[J]. Soybean Science, 2012,31(4):546-551. | |
[9] | 王慧梅, 王延兵, 祖元刚, 等. 干旱胁迫下黄檗幼苗cDNA消减文库的构建和分析[J]. 生物工程学报, 2008,24(2):198-202. |
Wang HM, Wang YB, Zu YG, et al. Construction and analysis of subtractive cDNA library of Phellodendron amurense under drought stress[J]. Chinese Journal Biotechnology, 2008,24(2):198-202. | |
[10] |
Kerk D, Bulgrien J, Smith DW, et al. Arabidopsis proteins containing similarity to the universal stress protein domain of bacteria[J]. Plant Physiol, 2003,131(3):1209-1219.
doi: 10.1104/pp.102.016006 URL pmid: 12644671 |
[11] |
Chou MX, Wei XY, Chen DS, et al. A novel nodule enhanced gene encoding a putative universal stress protein from Astragalus sinicus[J]. J Plant Physiol, 2007,164(6):764-772.
doi: 10.1016/j.jplph.2006.05.009 URL pmid: 16884822 |
[12] | 陈莹, 陈锡, 蔡一鸣, 等. 高羊茅春化基因FeVRN2的克隆及其在非生物胁迫下的表达[J]. 植物生理学报, 2018,54(6):1095-1102. |
Chen Y, Chen X, Cai YM, et al. Cloning and expression analysis of vernalization gene FaVRN2 under abiotic stress conditions in tall fescue[J]. Plant Physiology Journal, 2018,54(6):1095-1102. | |
[13] | 李小冬, 蔡璐, 李世歌, 等. 免预培养高效菊苣遗传转化方法[J]. 草业学报, 2016,25(10):124-131. |
Li XD, Cai L, Li SG, et al. An effective transformation method mediated by Agrobacterium in chicory[J]. Acta Prataculturae Sinica, 2016,25(10):124-131. | |
[14] | Nystrom T, Neidhardt FC. Isolation and properties of a mutant of Escherichia coli with an insertional inactivation of the uspa gene, which encodes a universal stress protein[J]. Bacteriol, 1993,175(39):49-56. |
[15] | Nystrom T, Neidhardt FC. Expression and role of the universal stress protein, UspA, of Escherichia coli during growth arrest[J]. Molecular Microbiology, 1994,11(5):37-44. |
[16] | 赵莘, 朱自果, 徐炎, 等. 中国野生华东葡萄抗白粉病相关基因(VpUSP)克隆及表达分析[J]. 果树学报, 2011,28(1):156-160. |
Zhao S, Zhu ZG, Xu Y, et al. Cloning and expression analysis of VpUSP gene in Chinese wild Vitis pseudoreticulata[J]. Journal of Fruit Science, 2011,28(1):156-160. | |
[17] |
Sauter M, Rzewuski G, Marwedel T, et al. The novel ethylene-regulated gene OsUsp1 from rice encodes a member of a plant protein family related to prokaryotic universal stress proteins[J]. Journal of Experimental Botany, 2002,53(379):2325-2331.
doi: 10.1093/jxb/erf096 URL pmid: 12432025 |
[18] | 刘丽, 徐兆师, 张瑞越, 等. 小麦胁迫相关基因W1的克隆及表达模式分析[J]. 植物遗传资源学报, 2008,9(4):423-427. |
Liu L, Xu ZS, Zhang RY, et al. Cloning and expression analysis of a stress related Gene W1 in Wheat[J]. Journal of Plant Genetic Resources, 2008,9(4):423-427. | |
[19] |
Bao GG, Zhuo CL, Qian CM, et al. Co-expression of NCED and ALO improves vitamin C level and tolerance to drought and chilling in transgenic tobacco and stylo plants[J]. Plant Biotechnology Journal, 2016,14:206-214.
doi: 10.1111/pbi.12374 URL pmid: 25865630 |
[20] |
Guttikonda SK, Valliyodan B, Neelakandan AK, et al. Over-expression of AtDREB1D transcription factor improves drought tolerance in soybean[J]. Mol Biol Rep, 2014,41:7995-8008.
URL pmid: 25192890 |
[21] |
Ksouri N, Jiménez S, Wells CE, et al. Transcriptional responses in root and leaf of Prunus persica under drought stress using RNA sequencing[J]. Frontiers in Plant Science, 2016,7:1715.
URL pmid: 27933070 |
[22] |
Chinnusamy V, Schumaker K, Zhu JK. Molecular genetic perspectives on cross-talk and specificity in abiotic stress signaling in plants[J]. J Exp Bot, 2004,55:225-236.
doi: 10.1093/jxb/erh005 URL pmid: 14673035 |
[23] |
Chi YH, Koo SS, Oh HT, et al. The physiological functions of universal stress proteins and their molecular mechanism to protect plants from environmental stresses[J]. Frontiers in Plant Science, 2019,10:750.
doi: 10.3389/fpls.2019.00750 URL pmid: 31231414 |
[24] |
Loukehaich R, Wang TT, Ouyang B, et al. SpUSP, an annexin-interacting universal stress protein, enhances drought tolerance in tomato[J]. J Exp Bot, 2012,63:5593-5606.
URL pmid: 22915741 |
[25] |
Pushpika U, Jha RK, Dinkar S, et al. Overexpression of a cytosolic abiotic stress responsive universal stress protein(SbUSP)mitigates salt and osmotic stress in transgenic tobacco plants[J]. Frontiers in Plant Science, 2016,7:518.
doi: 10.3389/fpls.2016.00518 URL pmid: 27148338 |
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