剑麻AhMADS8937基因酵母双杂交诱饵载体的构建、毒性和自激活检测  

Construction Toxicity Detection and Autoactivation of a Yeast Two-hybrid Bait Vector Containing a Sisal A hMA DS8937 Gene

作　　者：杨茜 徐蕾 汪询 杨子平[2] 周文钊[2] YangXi;Xu Lei;Wang Xun;Yang Ziping;Zhou Wenzhao(College of Horticulture,Hainan University,Haikou,570208;Zhanjiang Key Laboratory of Tropical Crop Genetic Improvement,South Subtropical Crops Research Institute,Chinese Academy of Tropical Agricultural Sciences,Zhanjiang,524091;College of Tropical Crops,Yunnan Agricultural University,Kunming,650100)

机构地区：[1]海南大学园艺学院,海口570208 [2]中国热带农业科学院南亚热带作物研究所,湛江市热带作物遗传改良重点实验室,湛江524091 [3]云南农业大学热带作物学院,昆明650100

出　　处：《分子植物育种》2024年第18期5994-5999,共6页Molecular Plant Breeding

基　　金：国家现代农业产业技术体系项目(CARS-16);国家自然科学基金青年基金项目(No.31801679);广东省甘蔗剑麻产业技术体系创新团队项目(No.2019KJ104-03)共同资助。

摘　　要：MADS-box是植物重要的转录调控因子,参与种子休眠和萌发、根系生长、花器官发育、果实成熟等营养生长和生殖生长过程。前期研究结果表明AhMADS8937可能参与调控剑麻株芽的发育。鉴于MADS-box蛋白能以复合体的形式调控植物发育,因此为筛选剑麻(Agave hybrid No.11648)中与AhMADS8937蛋白相互作用的蛋白,本研究构建了pGBKT7-AhMADS8937诱饵表达载体,酶切验证和测序结果表明载体构建成功。将重组载体转化Y2HGold酵母感受态,检测其对酵母菌株是否有毒性,以及是否存在自激活。结果表明pGBKT7-Ah MADS8937诱饵载体对Y2HGold酵母菌无毒性但存在自激活,5 mmol/L的3-氨基1,2,4-三唑(3-Amino-1,2,4-triazole,3-AT)可抑制其自激活。本研究成功构建AhMADS8937酵母双杂交诱饵载体,为筛选与AhMADS8937相互作用的蛋白,解析剑麻株芽发育的分子调控网络打下基础。MADS-box is an important transcription regulator in plants,which is involved in seed dormancy and germination,root growth,flower organ development,fruit maturation and other vegetative and reproductive growth processes.The results of previous studies indicated that AhMA DS8937 may be involved in regulating the development of bulbil of sisal.In view of the fact that MADS-box protein can regulate plant development in the form of complex,in order to screen the proteins interacting with AhMADS8937 protein in sisal(Agave hybrid No.11648),the bait of pGBKT7-AhMADS8937 vector was constructed.The restriction enzyme analysis and DNA sequencing showed that the vector was successfully constructed.The recombinant vector was transformed into Y2HGold yeast to detect its toxicity and autoactivation.The result showed that the bait vector had no toxicity to Y2HGold.However,it had autoactivation,which was inhibited by 5 mmol/L 3-AT.The study successfully constructed the bait vector of AhMADS8937 for yeast two-hybrid system,which laid a foundation for screening the proteins interacting with AhMADS8937.

关 键 词：剑麻 AhMADS8937 酵母双杂交 诱饵载体 毒性和自激活性 

分 类 号：S563.8[农业科学—作物学]