魔芋热处理响应基因AaHSP70启动子的克隆和功能分析  

Cloning and Functional Analysis of a Heat Treatment Response Factor Gene AaHSP70 in Amorphophallus

作　　者：王映红 张楠 岳振宇 刘海利 牛义 WANG Yinghong;ZHANG Nan;YUE Zhenyu;LIU Haili;NIU Yi(Collage of Horticulture and Landscape Architecture,Southwest University/Key Laboratory of Horticulture Science for Southern Mountainous Regions,Ministry of Education,Chongqing 400715,China)

机构地区：[1]西南大学园艺园林学院/南方山地园艺学教育部重点实验室,重庆400715

出　　处：《西南大学学报（自然科学版）》2023年第2期66-75,共10页Journal of Southwest University(Natural Science Edition)

基　　金：重庆市技术创新与应用发展专项(cstc2019jscx-gksbX0159,cstc2019jscx-msxmX0404);西南大学石柱创新基金项目(SZ201902)。

摘　　要：以白魔芋(Amorphophallus albus)为试材,利用同源克隆法获得热处理响应基因AaHSP70.qRT-PCR结果表明:AaHSP70基因对热胁迫较敏感,41℃处理能够诱导AaHSP70在白魔芋不同器官中表达,并在1 h时表达丰度最高;利用FPNI-PCR法通过3轮步移扩增得到1062 bp的AaHSP70启动子序列.生物信息学分析表明:AaHSP70含有热胁迫响应元件HSE及多种与植物逆境应答相关的顺式作用元件.酵母单杂交试验结果和双荧光素酶系统均显示:AaHSP70启动子可与AaHSFA1,AaHSFA2c相互作用.为进一步分析AaHSP70启动子的功能,构建该启动子与GUS基因融合的植物表达载体,利用农杆菌介导法转入拟南芥,热处理后对转基因拟南芥进行GUS组织化学染色鉴定,结果显示其在根、茎、叶中均可表达,因此推测AaHSP70可能在白魔芋抗外界逆境特别是热胁迫中起着重要的作用.The heat treatment response gene AaHSP70 was obtained by homologous cloning from Amorphophallus albus. The results of qRT-PCR showed that the AaHSP70 gene was more sensitive to heat stress. Heat treatment at 41 ℃ can induce the expression of AaHSP70 in different organs of Amorphophallus, with the highest expression apeared within 1 h. The 1 062 bp of AaHSP70 promoter sequence was obtained by FPNI-PCR method. Bioinformatic analysis showed that AaHSP70 contains heat stress response element HSE and a variety of cis-acting elements related to plant stress response. The results of yeast one-hybrid assay and the dual luciferase system showed that the promoter of AaHSP70 could interact with AaHSFA1 and AaHSFA2 c. To further analyze the function of the AaHSP70 promoter, a plant expression vector fused with the promoter and GUS gene was constructed and transferred into Arabidopsis by Agrobacterium-mediated method. After heat treatment, the transgenic Arabidopsis was identified by GUS histochemical staining. The results showed that AaHSP70 expressed in roots, stems and leaves. Therefore, it could be speculated that AaHSP70 may play an important role in the resistance of Amorphophallus to external adversity, especially heat stress.

关 键 词：白魔芋 AaHSP70 荧光定量 启动子 

分 类 号：S632.3[农业科学—蔬菜学] Q943.2[农业科学—园艺学]