月季切花乙烯受体RhETR3互作蛋白RhTSPO1的筛选和鉴定

园艺学报 ›› 2014, Vol. 41 ›› Issue (6): 1157-1166.

月季切花乙烯受体RhETR3互作蛋白RhTSPO1的筛选和鉴定

杨若韵1，陈雯1，薛璟祺2，田佶1，张帅1，朱春彦1，高俊平1，马男1,*

（1中国农业大学观赏园艺与园林系，北京 100193；2中国农业科学院蔬菜花卉研究所，北京 100081）

出版日期:2014-06-25 发布日期:2014-06-25

Screening and Identification of RhTSPO1，an Interacting Protein of Ethylene Receptor RhETR3 in Cut Roses

YANG Ruo-yun1，CHEN Wen1，XUE Jing-qi2，TIAN Ji1，ZHANG Shuai1，ZHU Chun-yan1，GAO Jun-ping1，and MA Nan1,*

（1Department of Ornamental Horticulture and Landscape Architecture，China Agricultural University，Beijing 100193，China；2Institute of Vegetables and Flowers，Chinese Academy of Agricultural Sciences，Beijing 100081，China）

Online:2014-06-25 Published:2014-06-25

摘要/Abstract

摘要： 利用已构建的月季分裂泛素酵母双杂文库，以月季乙烯受体蛋白RhETR3为诱饵，筛选能够与RhETR3结合的互作蛋白。经酵母回转验证共得到26个阳性克隆。在候选蛋白中发现一个TspO/MBR蛋白家族的成员，命名为RhTSPO1。通过RACE分离了RhTSPO1基因全长，共777 bp，开放阅读框（ORF）为579 bp，编码192个氨基酸。在酵母中，利用酵母生长和β–半乳糖苷酶活性（β-gal）验证了RhETR3和RhTSPO1之间确实存在互作。将RhETR3-GFP和RhTSPO1-mCherry融合蛋白共转入烟草，激光共聚焦显微镜观察发现RhTSPO1和RhETR3蛋白共同定位于内质网上。月季花朵自然开放过程中，RhTSPO1在花瓣中的表达呈现先增强后减弱的趋势，与开放和衰老过程紧密联系，表明RhTSPO1可能参与了花朵开放过程。本试验结果表明乙烯受体蛋白水平的互作调节可能也参与月季花朵开放和衰老的调节。

关键词: 月季, 花朵开放, 乙烯受体, 蛋白互作

Abstract: It is known that RhETR3 plays an important role in ethylene-regulated flower opening of roses（Rosa hybrida）. Here，RhETR3 was used as a bait to screen its potential interacting proteins in rose petals by using split ubiquitin yeast two-hybrid system. Of 26 positive clones which were identified and confirmed by retransformation in yeast，a TSPO/MBR protein was isolated by RACE and was named as RhTSPO1. The full length of RhTSPO1 was 777 bp which contained a 579 bp open reading frame，encoding 192 amino acids residues. The interaction between RhETR3 and RhTSPO1 was confirmed by β-galactosidase activity test in yeast. In addition，RhTSPO1 was fused with a mCherry fluorescent protein. The resultant RhTSPO1-mCherry fusion protein was used to conduct the fluorescence co-localization with RhETR3-GFP. The results indicated that these two proteins both located in the endoplasmic reticulum. The expression of RhTSPO1 increased as flower opening in the petals of cut roses，while it started to decrease when flower reached full opening，implying that RhTSPO1 may be involved in the flower opening process.

Key words: cut rose, flower opening, ethylene receptor, protein interaction

中图分类号:

S 685.12

杨若韵陈雯薛璟祺田佶张帅朱春彦高俊平马男. 月季切花乙烯受体RhETR3互作蛋白RhTSPO1的筛选和鉴定[J]. 园艺学报, 2014, 41(6): 1157-1166.

YANG Ruo-yun1，CHEN Wen1，XUE Jing-qi2，TIAN Ji1，ZHANG Shuai1，ZHU Chun-yan1，GAO Jun-ping1，and MA Nan1,*. Screening and Identification of RhTSPO1，an Interacting Protein of Ethylene Receptor RhETR3 in Cut Roses[J]. ACTA HORTICULTURAE SINICA, 2014, 41(6): 1157-1166.

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