| 条斑紫菜酵母双杂交文库的构建及PyMAPK5互作蛋白的筛选 |
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| 引用本文： | 董道英,孔凡娜,崔正彩,孙斌.条斑紫菜酵母双杂交文库的构建及PyMAPK5互作蛋白的筛选[J].海洋与湖沼,2019,50(6):1271-1280. |
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| 作者姓名： | 董道英 孔凡娜 崔正彩 孙斌 |
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| 作者单位： | 中国海洋大学海洋生命学院海洋生物遗传学与育种教育部重点实验室 青岛 266003 |
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| 基金项目： | 国家自然科学基金项目，31672641号。 |
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| 摘 要： | 为进一步研究条斑紫菜促分裂原活化激酶家族PyMAPK5的下游互作蛋白,理解其生物学功能,本研究通过酵母双杂交的方法进行其相互作用蛋白的筛选。提取不同温度和失水逆境胁迫下的RNA,利用Invitrogen体系构建条斑紫菜酵母双杂交cDNA文库,其库容为1.44×107CFU,重组率为91.8%。以pGBKT7-PyMAPK5为诱饵蛋白载体,利用共转化方法,从文库中筛选得到26个与PyMAPK5互作的候选蛋白。候选蛋白集中在光系统II相关蛋白、捕光蛋白、微管蛋白、ATP酶、GTP结合蛋白及假设蛋白等。微管蛋白、捕光蛋白、光系统II蛋白一对一验证结果为阳性,表明在酵母体内存在互作。本研究为阐明条斑紫菜PyMAPK5与其互作蛋白的关系及解析PyMAPK5下游作用机制奠定了基础。
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| 关 键 词： | PyMAPK5 条斑紫菜 酵母双杂交 互作蛋白 逆境胁迫 |
| 收稿时间： | 2019/4/5 0:00:00 |
| 修稿时间： | 2019/5/27 0:00:00 |
CONSTRUCTION OF A TWO-HYBRID LIBRARY OF PYROPIA YEZOENSIS AND SCREENING OF PYMAPK5 INTERACTION PROTEIN |
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| DONG Dao-Ying,KONG Fan-N,CUI Zheng-Cai and SUN Bin.CONSTRUCTION OF A TWO-HYBRID LIBRARY OF PYROPIA YEZOENSIS AND SCREENING OF PYMAPK5 INTERACTION PROTEIN[J].Oceanologia Et Limnologia Sinica,2019,50(6):1271-1280. |
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| Authors: | DONG Dao-Ying KONG Fan-N CUI Zheng-Cai and SUN Bin |
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| Institution: | The Key Laboratory of Marine Genetics and Breeding, Ministry of Education, Ocean University of China, Qingdao 266003, China,The Key Laboratory of Marine Genetics and Breeding, Ministry of Education, Ocean University of China, Qingdao 266003, China,The Key Laboratory of Marine Genetics and Breeding, Ministry of Education, Ocean University of China, Qingdao 266003, China and The Key Laboratory of Marine Genetics and Breeding, Ministry of Education, Ocean University of China, Qingdao 266003, China |
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| Abstract: | To understand the downstream interacting proteins of mitogen-activated kinase family PyMAPK5 and its biological functions in Pyropia yezoensis, we screened the interacting proteins of PyMAPK5 by yeast two-hybrid. Total RNA of P. yezoensis under a different temperature and water stress was used to construct the yeast cDNA library with the Invitrogen system. The resulted storage capacity was 1.44×107CFU, and the recombination rate was 91.8%. In addition, we screened the total 26 candidate proteins interacting with PyMAPK5 by co-transformation method. The candidate proteins were focused on photosystem II related proteins, light-harvesting proteins, tubulin, ATPase, GTP-binding proteins, and hypothetical proteins. At last, we one-to-one verified the reliabilities of three candidate proteins (including tubulin, light-harvesting protein, and photosystem II protein). This study may help clarify the relationship between PyMAPK5 and its interaction proteins and lay a foundation for the analysis of downstream mechanism of PyMAPK5. |
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| Keywords: | PyMAPK5 Pyropia yezoensis yeast two-hybrid interacting proteins adversity stress |
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