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[1]武广珩** 傅仙玉.利用CRISPR/Cas9技术编辑水稻负调控抗病基因OsEDR1及功能分析*[J].应用与环境生物学报,2020,26(02):1-10.[doi:10.19675/j.cnki.1006-687x.2019.05007]
 WU Guangheng,**,FU Xianyu.Editing rice negative regulation resistance gene OsEDR1 by CRISPR/Cas9 and its function analysis*[J].Chinese Journal of Applied & Environmental Biology,2020,26(02):1-10.[doi:10.19675/j.cnki.1006-687x.2019.05007]
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利用CRISPR/Cas9技术编辑水稻负调控抗病基因OsEDR1及功能分析*()
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《应用与环境生物学报》[ISSN:1006-687X/CN:51-1482/Q]

卷:
26卷
期数:
2020年02期
页码:
1-10
栏目:
研究论文
出版日期:
2020-03-20

文章信息/Info

Title:
Editing rice negative regulation resistance gene OsEDR1 by CRISPR/Cas9 and its function analysis*
作者:
武广珩12** 傅仙玉3
1福建省生态产业绿色技术重点实验室 武夷山 354300
2武夷学院生态与资源工程学院 武夷山 354300
3武夷学院茶与食品学院 武夷山 354300
Author(s):
WU Guangheng1 2** FU Xianyu3
1 Provincial Key Laboratory of Eco-Industrial Green Technology, Wuyishan 354300, China
2 College of Ecology and Resources Engineering, Wuyi University, Wuyishan 354300, China 3 College of Tea and Food Science, Wuyi University, Wuyishan 354300, China
关键词:
EDR1蛋白激酶基因编辑过表达白叶枯
Keywords:
EDR1 protein kinase gene editing over-expression bacterial blight
DOI:
10.19675/j.cnki.1006-687x.2019.05007
摘要:
拟南芥EDR1(Enhanced Disease Resistance1),一个Raf样MAPKKK蛋白激酶,通过级联蛋白激酶途径MKK4/MKK5–MPK3/MPK6负调控水杨酸诱导的免疫防卫反应。为了研究水稻OsEDR1基因的功能,我们利用CRISPR/Cas9 技术创制了OsEDR1的功能缺失突变体的同时,构建了过表达载体pTCK303-Ubi-OsEDR1转化水稻,获得转基因植株,最后接种白叶枯病菌。此外,我们也构建了pBI1.4t-35S-OsEDR1表达载体并转化拟南芥,分析了转基因植株的白粉菌抗病性。结果表明利用CRISPR/Cas9技术创制了单碱基插入的OsEDR1功能缺失突变体;白叶枯病原菌Xoo PXO99的接种实验表明,该功能缺失突变体较日本晴相比,病斑长度减少约50%,增强了水稻对Xoo PXO99的抗性。定量PCR和接种实验结果显示在日本晴中过量表达OsEDR1增强了水稻对Xoo PXO86的感病性。在拟南芥edr1突变体中异源表达OsEDR1无法抑制edr1突变体对白粉菌的抗病和白粉菌诱导的细胞死亡。综上所述,OsEDR1负调控水稻对白叶枯的抗病反应和自发的细胞死亡,但可能与拟南芥所依赖的防御反应信号通路不同。为下一步深入研究EDR1基因的作用机制和在水稻育种中的应用提供帮助。(图4 表1 参29)
Abstract:
EDR1(Enhanced Disease Resistance1) in Arabidopsis thaliana is a Raf-like MAPKK protein kinase that negatively regulates salicylic acid-induced defense response and controls plant immune defense response through MKK4/MKK5–MPK3/MPK6. In order to study the function of OsEDR1 gene in rice, we developed a loss-of-function mutant of OsEDR1 using CRISPR/Cas9 technology. Over-expression vectors pTCK303-Ubi-OsEDR1 and pBI1.4t-35S-OsEDR1 were constructed to transform rice and Arabidopsis to obtain transgenic plants. Finally, bacterial blight and powdery mildew were inoculated to analyze the resistance. The results showed that CRISPR/Cas9 technology created loss-of-function mutant of OsEDR1 with single base insertion. The inoculation experiment of Xoo PXO99 showed that the length of lesion of loss-of-function mutant was reduced by about 50% compared with that of Nipponbare, which enhanced resistance of rice to Xoo PXO99. Quantitative PCR and inoculation experiment results showed that over-expression of OsEDR1 in Nipponbare could enhance susceptibility to Xoo PXO86. The heterologous expression of OsEDR1 in Arabidopsis edr1 mutant could not compromise resistance of edr1 mutant to powdery mildew and cell death induced by powdery mildew. In conclusion, OsEDR1 negatively regulates resistance to bacterial blight and spontaneous cell death in rice, but it may be different from defense signaling in Arabidopsis. These results will be helpful to further study the mechanism of EDR1 gene and its application in rice breeding.

参考文献/References:


备注/Memo

备注/Memo:
收稿日期 Received: 2019-05-07 接受日期 Accepted: 2019-07-19
*福建省高校杰出青年科研人才培育计划(闽科教[2016]23号)、福建省自然科学基金青年项目(2016J05086)、武夷学院引进人才科研启动项目(YJ201503)资助
**通讯作者 Corresponding author (E-mail: wguangheng@163.com)
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更新日期/Last Update: 2019-07-30