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应用酵母单杂交系统分离水稻蜡质基因5^调控区结合蛋白的基因

陈丽, 邢彦彦, 张景六, 王宗阳, 洪孟民

中科院上海植物生理研究所.上海 200032

摘要

在水稻蜡质基因5^上游区中一段31bp核苷酸序列能与水稻未成熟种子核蛋白特异结合。为了克隆这一核蛋白基因,以此31bp序列构建成“鱼饵”质粒,从水稻cDNA文库中筛选到13个阳性克隆。根据这些阳性克隆中插入cDNA片段的相互杂交结果,对这些克隆进行了分组。

关键词： 水稻; 蜡质基因; 结合蛋白; 酵母单杂交系统

Isolation of the genes encording DNA binding protein interacting with 5^ upstream region of the rice waxy gene by using yeast one-hybrid system

Chen Li, Xing Yanyan, Zhang Jingliu, Wang Zongyang, Hong Mengmin

Shanghai Institute of Plant Physiology,Chinese Academy of Sciences.Shanghai 200032

Abstract

A 31 bp DNA fragment within the 5^ upstream region of rice waxy gene was used as a bait in a yeast one-hybrid system to isolate the genes encoding trans-acting factors regulating rice waxy gene expression. This yeast one-hybrid system consists of two plasmids: the p178 plasmid consisted of two copies of 31 bp oligonucleotids adjacent to the mini-promoter Pcyc1 directing lacZ gene expression, in the second pPC86 plasmid, a rice cDNA library was ligated to a gene encoding GAL4 transactivating domain as a translational fusion to form a rice cDNA-GAL4 library. A yeast strain harboured the p178 plasmid was then transformed with the pPC86 plasmid containing cDNA-GAL4 library, and grown on medium containing X-gal to select blue colonies. After several round selections, thirteen positive clones were obtained. These thirteen cDNA clones can be divided into four groups according to their homology by Southern blot analysis.

Key words: Rice;Waxy gene;Binding protein;Yeast one-hybrid system

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引用本文：

陈丽, 邢彦彦, 张景六, 王宗阳, 洪孟民. 应用酵母单杂交系统分离水稻蜡质基因5^调控区结合蛋白的基因[J]. 生理学报 1999; 51 (2): .

Chen Li, Xing Yanyan, Zhang Jingliu, Wang Zongyang, Hong Mengmin. Isolation of the genes encording DNA binding protein interacting with 5^ upstream region of the rice waxy gene by using yeast one-hybrid system. Acta Physiol Sin 1999; 51 (2): (in Chinese with English abstract).