OsRUS2.1酵母双杂交猎物载体的构建及其细胞毒性和自激活作用检测

潘家强<sup>1</sup>; $2<sup>1</sup>; <sup>2</sup>*

引用本文:	潘家强，侯学文，*.OsRUS2.1酵母双杂交猎物载体的构建及其细胞毒性和自激活作用检测[J].广西植物,2013,(1):76-81.[点击复制]
	PAN Jia Qiang， HOU Xue Wen，*.Construction of OsRUS 2.1 yeast two hybrid prey vectors and detection of their toxicity and self activated activity[J].Guihaia,2013,(1):76-81.[点击复制]

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OsRUS2.1酵母双杂交猎物载体的构建及其细胞毒性和自激活作用检测
潘家强¹，侯学文¹，²*
1.华南农业大学生命科学学院分子植物生理研究室，广州 510642;2.华南农业大学 生命科学学院植物功能基因组与生物技术重点实验室，广州 510642

摘要:

采用PCR技术扩增水稻根UV B敏感基因2.1（ROOT UV B SENSITIVE 2.1，RUS2.1）四个不同片段[OsRUS2.1(1 1317)，OsRUS2.1(1 138)，OsRUS2.1(139 879)，OsRUS2.1(880 1317)]，连接到T载体pMD18 T Simple上，测序无误后分别亚克隆到猎物表达载体pGADT7上。结果表明：四个OsRUS2.1基因片段的表达载体构建成功，读码框正确；分别转化这四个猎物表达载体于酵母感受态细胞Y187中，用LacZ、MEL1活性检测法和营养缺陷型培养基SD Leu DO培养法进行自激活检测和毒性检测，结果表明构建的四个OsRUS2.1不同片段的猎物表达载体对酵母菌株Y187均没有转录激活活性和毒害作用，可用于后续研究。

关键词: 水稻根对UVB敏感基因2.1 猎物载体酵母双杂交自激活毒性检测

DOI：

分类号:Q782

基金项目:

Construction of OsRUS 2.1 yeast two hybrid prey vectors and detection of their toxicity and self activated activity

PAN JiaQiang¹， HOU XueWen¹，²*

1.Lab of Molecular Plant Physiology， College of Life Sciences， South China Agricultural University， Guangzhou 510642， China;2.Key Laboratory of Plant Functional Genomics and Biotechnology， College of Life Sciences， South China Agricultural University， Guangzhou 510642， China 1.Lab of Molecular Plant Physiology， College of Life Sciences， South China Agricultural University， Guangzhou 510642， China； 2.Key Laboratory of Plant Functional Genomics and Biotechnology， College of Life Sciences， South China Agricultural University， Guangzhou 510642， China

Abstract:

Four fragments of rice(Oryza sativa)ROOT UV B SENSITIVE 2.1(OsRUS2.1)，OsRUS2.1(1 1317)，OsRUS2.1(1 138)，OsRUS2.1(139 879)，OsRUS2.1(880 1317)，were amplified by PCR from cloned OsRUS2.1 plasmid，and were ligated with pMD18 T simple vector，then transformed to E.coli TOP10 competent cell. The positive clones were selected and sequenced. The confirmed fragments were subcloned to prey vector pGADT7. The four constructed pGADT7 prey vectors were further confirmed by enzyme digestion and sequencing. The confirmed 4 types of pGADT7 prey vectors were transformed to Y187 yeast competent cells. The self activation and toxicity of the plasmids to host yeast Y187 were detected by LacZ and MEL1 activity assays and culturing in auxotroph medium SD Leu DO． Results showed that the four constructed plasmids had no self transcriptional activity and not toxicity to yeast strain Y187，and they could be used in the following yeast two hybrid experiments．

Key words: rice ROOT UV B SENSITIVE 2.1(OsRUS2.1) prey vector yeast two hybrid self activated transcriptional activity toxicity detection