欢迎访问作物学报，今天是
作物学报

作物学报 ›› 2020, Vol. 46 ›› Issue (8): 1217-1224.doi: 10.3724/SP.J.1006.2020.92060

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

OsRPK1基因过表达和RNA干涉对水稻苗期耐盐性的影响

李晶岚,陈鑫欣,石翠翠,刘方惠,孙静,葛荣朝()   

  1. 河北师范大学生命科学学院, 河北石家庄 050024
  • 收稿日期:2019-11-25 接受日期:2020-03-24 出版日期:2020-08-12 网络出版日期:2020-04-21
  • 通讯作者: 葛荣朝
  • 作者简介:李晶岚, E-mail: 83494429@qq.com|陈鑫欣, E-mail: 1046433763@qq.com
  • 基金资助:
    国家自然科学基金项目(30900104);北省自然科学基金项目(C2016205158)

Effects of OsRPK1 gene overexpression and RNAi on the salt-tolerance at seedling stage in rice

LI Jing-Lan,CHEN Xin-Xin,SHI Cui-Cui,LIU Fang-Hui,SUN Jing,GE Rong-Chao()   

  1. College of Life Science, Hebei Normal University, Shijiazhuang 050024, Hebei, China
  • Received:2019-11-25 Accepted:2020-03-24 Published:2020-08-12 Published online:2020-04-21
  • Contact: Rong-Chao GE
  • Supported by:
    National Natural Science Foundation of China(30900104);Hebei Provincial Natural Science Foundation(C2016205158)

RichHTML

31

PDF

965

摘要：

水稻OsRPK1基因属于富含亮氨酸重复序列的类受体蛋白激酶, 在盐胁迫下其表达量暂时升高后出现明显下降。前期研究表明, OsRPK1基因过表达会造成拟南芥非生物胁迫耐受性明显降低。本研究对OsRPK1基因在水稻中实现过表达和RNA干涉, 进而对OsRPK1的抗逆性功能加以探究。结果表明, OsRPK1基因表达量增加时, 水稻幼苗表现为耐盐性下降; RNA干涉则使水稻幼苗表现为耐盐性增加。盐胁迫后OsRPK1过表达植株脯氨酸含量低于野生型、MDA含量和相对电导率显著高于野生型, 而OsRPK1-RNAi水稻植株的脯氨酸含量显著高于野生型、MDA含量和相对电导率显著低于野生型。因此, OsRPK1基因的表达量对水稻耐盐性具有明显影响, 脯氨酸含量及细胞质膜受破损程度的改变可能是造成转基因水稻耐盐性变化的内在原因。本研究为进一步阐明OsRPK1基因的抗逆作用机制奠定了基础, 对于通过调整该基因表达改良水稻的耐盐性具有重要意义。

关键词: 水稻, OsRPK1基因, 过表达, RNA干涉, 生理指标

Abstract:

OsRPK1 is a leucine-rich repeat receptor-like protein kinase (LRR-RLK) gene in rice, and its expression level decreases evidently after temporarily increasing under salt stress. Previous studies have shown that the overexpression of OsRPK1 can significantly reduce the abiotic stress tolerance of Arabidopsis. In this study, OsRPK1 gene was overexpressed and RNA interfered in rice. The salt tolerance of the 35S:OsRPK1 rice seedlings was significantly lower than that of the control plants. The OsRPK1-RNAi rice seedlings exhibited higher salt tolerance than the wild-type plants. The proline content of OsRPK1 overexpressed plants was lower than that of the wild type, MDA content and relative conductivity were significantly higher than those of the wild type, while the proline content of OsRPK1-RNAi rice plants was significantly higher than those of wild type, MDA content and relative conductivity were significantly lower than those of wild type. The results of this study indicate that OsRPK1 plays an important role in salt tolerance of rice. The changes of proline content and the damage degree of cytoplasmic membrane may be the internal reasons for the salt-tolerance variations of the transgenic rice lines. This study lays a foundation for further elucidating the resistance mechanism of OsRPK1 gene, which is very important for cultivating salt-tolerance rice by adjusting the expression of OsRPK1 gene.

Key words: rice, OsRPK1 genes, overexpressed, RNA interference, physiological indexes

图1

OsRPK1基因在140 mmol L-1 NaCl胁迫的水稻中表达量的变化 差异显著性分析采用t-test方法, *P < 0.05、**P < 0.01、***P < 0.001。"

图2

35S:OsRPK1植株与OsRPK1-RNAi植株的qRT-PCR检测 差异显著性分析采用t-test方法, *P < 0.05、**P < 0.01、 ***P < 0.001。"

图3

OsRPK1过表达水稻在盐胁迫下的生长情况 A: NaCl胁迫前; B: 140 mmol L-1 NaCl胁迫6 d; C: 140 mmol L-1 NaCl胁迫后恢复9 d; D: 盐胁迫恢复培养后存活率统计。差异显著性分析采用t-test方法, *P < 0.05、**P < 0.01、***P < 0.001。"

图4

OsRPK1-RNAi水稻在胁迫下生长情况 A: NaCl胁迫前; B: 140 mmol L-1 NaCl胁迫10 d; C: 140 mmol L-1 NaCl胁迫后恢复7 d; D, E: 盐胁迫恢复培养后存活率统计。差异显著性分析采用t-test方法, *P < 0.05、**P < 0.01、***P < 0.001。"

图5

140 mmol L-1 NaCl盐胁迫后转基因水稻脯氨酸含量 差异显著性分析采用t-test方法, *P < 0.05、**P < 0.01、***P < 0.001。"

图6

140 mmol L-1 NaCl盐胁迫后转基因水稻株系中MDA含量 差异显著性分析采用t-test方法, *P < 0.05、**P < 0.01、***P<0.001。"

图7

转基因水稻的相对电导率 差异显著性分析采用t-test方法, *P < 0.05、**P < 0.01、***P < 0.001。"

[1] Liang W J, Ma X L, Wan P, Liu L Y. Plant salt-tolerance mechanism: a review. Biochem Biophy Res Commun, 2017,495:286-291.
[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(3):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(3):154-158 (in Chinese with English abstract).
[3] 张建峰, 张旭东, 周金星, 刘国华, 李冬雪. 世界盐碱地资源及其改良利用的基本措施. 水土保持研究, 2005,12(6):28-30.
Zhang J F, Zhang X D, Zhou J X, Liu G H, Li D X. World resources of saline soil and main amelioratior measures. Res Soil Water Conserv, 2005,12(6):28-30 (in Chinese with English abstract).
[4] Lee S K, Kim B G, Kwon T R, Jeong M J, Park S R, Lee J W, Byun M O, Kwon H B, Matthews B F, Hong C B, Park S C. Overexpression of the mitogen-activated protein kinase gene OsMAPK33 enhances sensitivity to salt stress in rice(Oryza sativa L.). J Biosci, 2011,36:139-151.
pmid: 21451255
[5] Xing H T, Guo P, Xia X L, Yin W L. PdERECTA, a leucine-rich repeat receptor-like kinase of poplar, confers enhanced water use efficiency in Arabidopsis. Planta, 2011,234:229-241.
pmid: 21399949
[6] Mondal T K, Rawal H C, Chowrasia S, Varshney D, Panda A K, Mazumdar A, Kaur H, Gaikwad K, Sharma T R, Singh N K. Draft genome sequence of first monocot-halophytic species Oryza coarctata reveals stress-specific genes. Sci Rep, 2018,8:13698-13708.
pmid: 30209320
[7] 牛吉山. 植物和小麦蛋白激酶的研究现状. 西北植物学报, 2003,23:143-150.
Niu J S. Studies on plant and wheat protein kinases. Acta Bot Boreali-Occident Sin, 2003,23:143-150 (in Chinese with English abstract).
[8] Julie M S, John C W. Plant protein kinase families and signal transduction. Plant Physiol, 1995,108:451-457.
[9] Depuydt S, Rodriguez-Villalon A, Santuari L, Wyser-Rmili C, Ragni L, Hardtke C S. Suppression of Arabidopsis protophloem differentiation and root meristem growth by CLE45 requires the receptor-like kinase BAM3. Proc Natl Acad Sci USA, 2013,110:7074-7079.
doi: 10.1073/pnas.1222314110 pmid: 23569225
[10] Bryan A C, Obaidi A, Wierzba M, Tax F E. XYLEM INTERMIXED WITH PHLOEM1, a leucine-rich repeat receptor-like kinase required for stem growth and vascular development in Arabidopsis thaliana. Planta, 2012,235:111-122.
pmid: 21853254
[11] Burr C A, Leslie M E, Orlowski S K, Chen I, Wright C E, Daniels M J, Liljegren S J. CAST AWAY, a membrane-associated receptor-like kinase, inhibits organ abscission in Arabidopsis. Plant Physiol, 2011,156:1837-1850.
doi: 10.1104/pp.111.175224 pmid: 21628627
[12] Mizuno S, Osakabe Y, Maruyama K, Ito T, Osakabe K, Sato T, Shinozaki K, Yamaguchi-Shinozaki K. Receptor-like protein kinase 2 (RPK2) is a novel factor controlling anther development in Arabidopsis thaliana. Plant J, 2007,50:751-766.
pmid: 17419837
[13] Engelsdorf T, Hamann T. An update on receptor-like kinase involvement in the maintenance of plant cell wall integrity. Ann Bot, 2014,114:1339-1347.
doi: 10.1093/aob/mcu043 pmid: 24723447
[14] Wu Y, Zhou J M. Receptor-like kinases in plant innate immunity. J Integr Plant Biol, 2013,55:1271-1286.
doi: 10.1111/jipb.12123 pmid: 24308571
[15] Phillips S M, Dubery I A, Heerden H. Identification and molecular characterisation of a lectin receptor-like kinase (GHLECRK-2) from cotton. Plant Mol Biol Rep, 2013,31:9-20.
[16] Singh P, Kuo Y C, Mishra S, Tsai C H, Chien C C, Chen C W, Desclos-Theveniau M, Chu P W, Schulze B, Chinchilla D, Boller T, Zimmerli L. The lectin receptor kinase-VI.2 is required for priming and positively regulates Arabidopsis pattern-triggered immunity. Plant Cell, 2012,24:1256-1270.
pmid: 22427336
[17] Tanaka H, Osakabe Y, Katsura S, Mizuno S, Maruyama K, Kusakabe K, Mizoi J, Shinozaki K, Yamaguchi-Shinozaki K. Abiotic stress-inducible receptor-like kinases negatively control ABA signaling in Arabidopsis. Plant J: Cell Mol Biol, 2012,70:599-613.
[18] Hua D, Wang C, He J, Liao H, Duan Y, Zhu Z, Guo Y, Chen Z, Gong Z. A plasma membrane receptor kinase, GHR1, mediates abscisic acid- and hydrogen peroxide-regulated stomatal movement in Arabidopsis. Plant Cell, 2012,24:2546-2561.
doi: 10.1105/tpc.112.100107 pmid: 22730405
[19] Yuriko O, Shinji M, Hidenori T, Maruyama K, Osakabe K, Todaka D, Fujita Y, Kobayashi M, Shinozaki K, Yamaguchi- Shinozaki K. Overproduction of the membrane-bound receptor- like protein kinase1, RPK1, enhances abiotic stress tolerance in Arabidopsis. J Biol Chem, 2010,285:9190-9201.
doi: 10.1074/jbc.M109.051938 pmid: 20089852
[20] Yang T, Chaudhuri S, Yang L, Du L, Poovaiah B W. A calcium/calmodulin-regulated member of the receptor-like kinase family confers cold tolerance in plants. J Biol Chem, 2010,285:7119-7126.
doi: 10.1074/jbc.M109.035659 pmid: 20026608
[21] Sicilia A, Testa G, Santoro D F, Cosentino S L, Lo Piero A R. RNASeq analysis of giant cane reveals the leaf transcriptome dynamics under long-term salt stress. BMC Plant Biol, 2019,19:355-379.
doi: 10.1186/s12870-019-1964-y pmid: 31416418
[22] Song W Y, Wang G L, Chen L L, Kim H S, Pi L Y, Holsten T, Gardner J, Wang B, Zhai W X, Zhu L H, Fauquet C, Ronald P. A receptor kinase-like protein encoded by the rice disease resistance gene Xa21. Science, 1995,270:1804-1806.
doi: 10.1126/science.270.5243.1804 pmid: 8525370
[23] Kobe B, Deisenhofer J. The leucine-rich repeat: a versatile binding motif. Trends Biochem Sci, 1994,19:415-421.
doi: 10.1016/0968-0004(94)90090-6 pmid: 7817399
[24] Jones D A, Jones J D G. The role of leucine-rich repeat proteins in plant defences. Adv Bot Res, 1997,24:89-167.
[25] Hong S W, Jon J H, Kwak J M, Nam H G. Identification of a receptor-like protein kinase gene rapidly induced by abscisic acid, dehydration, high salt, and cold treatments in Arabidopsis thaliana. Plant Physiol, 1997,113:1203-1212.
doi: 10.1104/pp.113.4.1203 pmid: 9112773
[26] Osakabe Y, Maruyama K, Seki M, Satou M, Shinozaki K, Yamaguchi-Shinozaki K. Leucine-rich repeat receptor-like kinase1 is a key membrane-bound regulator of abscisic acid early signaling in Arabidopsis. Plant Cell, 2005,17:1105-1119.
doi: 10.1105/tpc.104.027474 pmid: 15772289
[27] Shi C C, Feng C C, Yang M M, Li J L, Li X X, Zhao B C, Huang Z J, Ge R C. Overexpression of the receptor-like protein kinase genes AtRPK1 and OsRPK1 reduces the salt tolerance of Arabidopsis thaliana. Plant Sci, 2014, 217-218:63-70.
doi: 10.1016/j.plantsci.2013.12.002 pmid: 24467897
[28] Wang J, Li C, Yao X, Liu S H, Zhang P Y, Chen K S. The Antarctic moss leucine-rich repeat receptor-like kinase (PnLRR-RLK2) functions in salinity and drought stress adaptation. Polar Biol, 2018,41:353-364.
[29] Sun X, Sun M, Luo X, Ding X, Cai H, Bai X, Liu X, Zhu Y. A Glycine soja, ABA-responsive receptor-like cytoplasmic kinase, GsRLCK, positively controls plant tolerance to salt and drought stresses. Planta, 2013,237:1527-1545.
doi: 10.1007/s00425-013-1864-6 pmid: 23494614
[30] An S H, Choi H W, Hwang I S, Hong J K, Hwang B K. A novel pepper membrane-located receptor-like protein gene CaMRP1 is required for disease susceptibility, methyl jasmonate insensitivity and salt tolerance. Plant Mol Biol, 2008,67:519-533.
doi: 10.1007/s11103-008-9337-1 pmid: 18427932
[31] Li C H, Wang G, Zhao J L, Zhang L Q, Ai L F, Han Y F, Sun D Y, Zhang S W, Sun Y. The receptor-like kinase SIT1 mediates salt sensitivity by activating MAPK3/6 and regulating ethylene homeostasis in rice. Plant Cell, 2014,26:2538-2553.
doi: 10.1105/tpc.114.125187
[32] Liu S H, Wang J, Chen K S, Zhang Z H, Zhang P Y. The L-type lectin receptor-like kinase(PnLecRLK1) from the Antarctic moss Pohlia nutans enhances chilling-stress tolerance and abscisic acid sensitivity in Arabidopsis. Plant Growth Regul, 81:409-418.
doi: 10.1007/s10725-016-0217-4
[1] 田甜, 陈丽娟, 何华勤. 基于Meta-QTL和RNA-seq的整合分析挖掘水稻抗稻瘟病候选基因[J]. 作物学报, 2022, 48(6): 1372-1388.
[2] 郑崇珂, 周冠华, 牛淑琳, 和亚男, 孙伟, 谢先芝. 水稻早衰突变体esl-H5的表型鉴定与基因定位[J]. 作物学报, 2022, 48(6): 1389-1400.
[3] 周文期, 强晓霞, 王森, 江静雯, 卫万荣. 水稻OsLPL2/PIR基因抗旱耐盐机制研究[J]. 作物学报, 2022, 48(6): 1401-1415.
[4] 郑小龙, 周菁清, 白杨, 邵雅芳, 章林平, 胡培松, 魏祥进. 粳稻不同穗部籽粒的淀粉与垩白品质差异及分子机制[J]. 作物学报, 2022, 48(6): 1425-1436.
[5] 颜佳倩, 顾逸彪, 薛张逸, 周天阳, 葛芊芊, 张耗, 刘立军, 王志琴, 顾骏飞, 杨建昌, 周振玲, 徐大勇. 耐盐性不同水稻品种对盐胁迫的响应差异及其机制[J]. 作物学报, 2022, 48(6): 1463-1475.
[6] 杨建昌, 李超卿, 江贻. 稻米氨基酸含量和组分及其调控[J]. 作物学报, 2022, 48(5): 1037-1050.
[7] 杨德卫, 王勋, 郑星星, 项信权, 崔海涛, 李生平, 唐定中. OsSAMS1在水稻稻瘟病抗性中的功能研究[J]. 作物学报, 2022, 48(5): 1119-1128.
[8] 朱峥, 王田幸子, 陈悦, 刘玉晴, 燕高伟, 徐珊, 马金姣, 窦世娟, 李莉云, 刘国振. 水稻转录因子WRKY68在Xa21介导的抗白叶枯病反应中发挥正调控作用[J]. 作物学报, 2022, 48(5): 1129-1140.
[9] 王小雷, 李炜星, 欧阳林娟, 徐杰, 陈小荣, 边建民, 胡丽芳, 彭小松, 贺晓鹏, 傅军如, 周大虎, 贺浩华, 孙晓棠, 朱昌兰. 基于染色体片段置换系群体检测水稻株型性状QTL[J]. 作物学报, 2022, 48(5): 1141-1151.
[10] 王泽, 周钦阳, 刘聪, 穆悦, 郭威, 丁艳锋, 二宫正士. 基于无人机和地面图像的田间水稻冠层参数估测与评价[J]. 作物学报, 2022, 48(5): 1248-1261.
[11] 陈悦, 孙明哲, 贾博为, 冷月, 孙晓丽. 水稻AP2/ERF转录因子参与逆境胁迫应答的分子机制研究进展[J]. 作物学报, 2022, 48(4): 781-790.
[12] 袁大双, 邓琬玉, 王珍, 彭茜, 张晓莉, 姚梦楠, 缪文杰, 朱冬鸣, 李加纳, 梁颖. 甘蓝型油菜BnMAPK2基因的克隆及功能分析[J]. 作物学报, 2022, 48(4): 840-850.
[13] 王吕, 崔月贞, 吴玉红, 郝兴顺, 张春辉, 王俊义, 刘怡欣, 李小刚, 秦宇航. 绿肥稻秆协同还田下氮肥减量的增产和培肥短期效应[J]. 作物学报, 2022, 48(4): 952-961.
[14] 巫燕飞, 胡琴, 周棋, 杜雪竹, 盛锋. 水稻延伸因子复合体家族基因鉴定及非生物胁迫诱导表达模式分析[J]. 作物学报, 2022, 48(3): 644-655.
[15] 陈云, 李思宇, 朱安, 刘昆, 张亚军, 张耗, 顾骏飞, 张伟杨, 刘立军, 杨建昌. 播种量和穗肥施氮量对优质食味直播水稻产量和品质的影响[J]. 作物学报, 2022, 48(3): 656-666.
Viewed
Full text


Abstract

Cited
  Shared   
  Discussed   
No Suggested Reading articles found!
京ICP备15008789号-2