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水稻LRR型受体蛋白激酶OsRPK1胞外区酵母双杂交诱饵载体构建
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作者 朱晓南 《安徽农业科学》 CAS 2015年第31期45-46,144,共3页
[目的]寻找水稻LRR型蛋白受体激酶(Leucine rich repeat receptor protein kinase)OsRPK1胞外结构域的互作蛋白。[方法]以日本晴水稻为研究材料,以水稻根部c DNA为模板通过PCR扩增方法得到OsRPK1胞外结构域基因编码片段OsRPK1-OD(969 bp... [目的]寻找水稻LRR型蛋白受体激酶(Leucine rich repeat receptor protein kinase)OsRPK1胞外结构域的互作蛋白。[方法]以日本晴水稻为研究材料,以水稻根部c DNA为模板通过PCR扩增方法得到OsRPK1胞外结构域基因编码片段OsRPK1-OD(969 bp)。将该片段插入酵母双杂交诱饵载体p GBKT7,构建重组诱饵载体p GBKT7-OsRPK1-OD。[结果]对pGBKT7-OsRPK1-OD进行毒性检测和转录自激活鉴定,酵母生长结果发现,含有pGBKT7-OsRPK1-OD载体的Y2H Gold细胞,在SD/-Trp、SD/-Trp/X-α-gal营养缺陷型固体培养基上生长出白色菌落,在SD/-Trp/X-α-gal/Ab A固体培养基上生长受到抑制;在SD/-Trp生长出含有pGBKT7-OsRPK1-OD载体的Y2H Gold酵母菌落大小与转化了空载的酵母菌落大小一致,显示插入的OsRPK1-OD对酵母生长无毒性作用。[结论]OsRPK1-OD无法参与激活转录酵母中报告基因HIS3,ADE2,MEL1和AUR1-C,该片段可以作为诱饵基因筛查水稻c DNA文库,寻找与之相互作用的蛋白质。 展开更多
关键词 水稻受体激酶 OsRPK 酵母双杂交 诱饵载体
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水稻富亮氨酸重复类受体蛋白激酶OsRPK1响应外源生长素的作用研究 被引量:2
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作者 邹禹 刘园园 +2 位作者 张培江 钱宝云 张炜 《江苏农业科学》 2019年第8期46-51,共6页
为明确水稻富亮氨酸重复类受体蛋白激酶OsRPK1响应外源生长素的作用机制,首先分析外源生长素2,4-D对OsRPK1转基因水稻根部表型的影响;其次,异源表达OsRPK1胞外富亮氨酸重复LRRs,检测H^3-IAA竞争结合融合蛋白GST-LRRs后的放射性活度以及... 为明确水稻富亮氨酸重复类受体蛋白激酶OsRPK1响应外源生长素的作用机制,首先分析外源生长素2,4-D对OsRPK1转基因水稻根部表型的影响;其次,异源表达OsRPK1胞外富亮氨酸重复LRRs,检测H^3-IAA竞争结合融合蛋白GST-LRRs后的放射性活度以及利用等温滴定量热(ITC)法分析IAA与融合蛋白GST-LRRs相互作用的微热量变化。结果表明,在正常生长条件下,OsRPK1过表达能够抑制水稻侧根的生长;0.01μmol/L 2,4-D处理4 d后发现,OsRPK1抑制表达植株侧根的数量比野生型多112%(P<0.01),而OsRPK1过表达植株侧根生长受到极显著抑制(P<0.001);0.1μmol/L 2,4-D处理10 d后,抑制表达植株不定根的数量相对于野生型多18.2%(P<0.05),而过表达植株不定根的生长受到显著抑制(P<0.01);大肠杆菌BL21(DE3)诱导表达获得了包涵体形式的GST-LRRs融合蛋白,通过复性、纯化获得可溶性融合蛋白;H^3-IAA竞争结合融合蛋白GST-LRRs以及IAA溶液滴定融合蛋白的微热量分析都表明OsRPK1与外源生长素未发生直接作用。 展开更多
关键词 水稻富亮氨酸重复类受体蛋白激酶OsRPK1 原核表达 外源生长素 融合蛋白GST-LRRs
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Screening of Extracellular Binding Proteins of Rice Receptor-like Kinase CR4 by the Yeast Two-hybrid 被引量:1
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作者 姚清国 李晓芹 +3 位作者 张文娜 周二鹏 王娟 王景翔 《Agricultural Science & Technology》 CAS 2010年第11期77-81,共5页
[Objective] The research aimed to find the extracellular binding proteins of CR4.[Method] The extracellular domain of OsCR4 was as the bait protein,and the yeast two-hybrid was used to screen cDNA library of seedling ... [Objective] The research aimed to find the extracellular binding proteins of CR4.[Method] The extracellular domain of OsCR4 was as the bait protein,and the yeast two-hybrid was used to screen cDNA library of seedling which was cultivated 14 d.[Result] A lot of proteins which included a peroxide B(D26484),a methionine thioredoxin reductase(ABF96078)and an unknown function protein were gained.[Conclusion] It provided the theory basis for studying the signal transduction mechanism of CR4. 展开更多
关键词 RICE Receptor-like kinase Extracellular binding protein Yeast two-hybrid
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A Xanthomonas oryzae pv. oryzae effector, Xop R, associates with receptor-like cytoplasmic kinases and suppresses PAMP-triggered stomatal closure 被引量:7
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作者 Shuangfeng Wang Jianhang Sun +3 位作者 Fenggui Fan Zhaoyun Tan Yanmin Zou Dongping Lu 《Science China(Life Sciences)》 SCIE CAS CSCD 2016年第9期897-905,共9页
Receptor-like kinases(RLKs) play important roles in plant immunity signaling; thus, many are hijacked by pathogen effectors to promote successful pathogenesis. Xanthomonas oryzae pv. oryzae(Xoo) is the causal agent of... Receptor-like kinases(RLKs) play important roles in plant immunity signaling; thus, many are hijacked by pathogen effectors to promote successful pathogenesis. Xanthomonas oryzae pv. oryzae(Xoo) is the causal agent of rice leaf blight disease. The strain PXO99 A has 18 non-TAL(transcription activation-like) effectors; however, their mechanisms of action and host target proteins remain largely unknown. Although the effector XopR from the Xoo strain MAFF311018 was shown to suppress PAMP-triggered immune responses in Arabidopsis, its target has not yet been identified. Here, we show that PXO99 A XopR interacts with BIK1 at the plasma membrane. BIK1 is a receptor-like cytoplasmic kinase(RLCK) belonging to the RLK family of proteins and mediates PAMP-triggered stomatal immunity. In turn, BIK1 phosphorylates XopR. Furthermore, XopR suppresses PAMP-triggered stomatal closure in transgenic Arabidopsis expressing XopR. In addition, XopR is able to associate with RLCKs other than BIK1. These results suggest that XopR likely suppresses plant immunity by targeting BIK1 and other RLCKs. 展开更多
关键词 XANTHOMONAS EFFECTOR XopR BIK1 immunity
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