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作物学报 ›› 2016, Vol. 42 ›› Issue (07): 1094-1099.doi: 10.3724/SP.J.1006.2016.01094

• 研究简报 • 上一篇    

花生AhDGAT2a基因启动子的克隆和功能验证

郑玲1,2,史灵敏1,3,田海莹1,2,单雷1,2,边斐1,郭峰1,宣宁1,万书波1,2,*,彭振英1,2,3,*   

  1. 1山东省农业科学院生物技术研究中心 / 山东省作物遗传改良与生态生理重点实验室,山东济南 250100; 2山东大学生命科学学院,山东济南 250100;3新疆农业大学农学院,新疆乌鲁木齐 830000
  • 收稿日期:2016-01-11 修回日期:2016-05-09 出版日期:2016-07-12 网络出版日期:2016-05-11
  • 通讯作者: 万书波, E-mail: wansb@saas.ac.cn, Tel: 0531-83178335; 彭振英, E-mail: pengzhenying2005@126.com
  • 基金资助:

    本研究由山东省自然科学基金项目(ZR2013CM036)和山东省良种工程项目(2014-2017)的资助。

Cloning and Functional Analysis of Peanut AhDGAT2a Promoter?

ZHENG Ling1,2,SHI Ling-Min1,3,TIAN Hai-Ying1,2,SHAN Lei1,2,BIAN Fei1,GUO Feng1, XUAN Ning1,WAN Shuo-Bo1,2,*,PENG Zhen-Ying1,2,3,*   

  1. 1 Biotechnology Research Center, Shandong Academy of Agricultural Sciences / Shandong Provincial Key Laboratory of Crop Genetic Improvement, Ecology and Physiology, Jinan 250100, China; 2 College of Life Science, Shandong University, Jinan 250100, China; 3 College of Agronomy, Xinjiang Agricultural University, Urumqi 830000, China
  • Received:2016-01-11 Revised:2016-05-09 Published:2016-07-12 Published online:2016-05-11
  • Contact: 万书波, E-mail: wansb@saas.ac.cn, Tel: 0531-83178335; 彭振英, E-mail: pengzhenying2005@126.com
  • Supported by:

    This study was supported by Natural Science Foundation of Shandong (ZR2013CM036) and Shandong Province Germplasm Innovation and Utilization Project (2014-2017).

摘要:

二酰甘油酰基转移酶(DGAT)是三酰甘油(TAG)合成途径的限速酶,对脂肪酸合成的调节具有关键作用。为了研究AhDGAT2a的表达调控,利用GenomeWalking方法从鲁花14基因组中克隆了AhDGAT2a上游5¢侧翼调控区1200 bp序列,即AhDGAT2a启动子(pAhDGAT2a)序列,并利用生物信息学软件分析其包含的调控元件,发现其含有多个TATA-box和CAAT-box、光调控元件、胁迫防御相关元件和激素响应元件。用pAhDGAT2a构建pAhDGAT2a:GUS植物表达载体并转化烟草品种SR1。利用组织染色法鉴定转基因烟草的GUS表达模式,发现在转基因烟草的各个器官均有GUS酶活,而在柱头、花药和幼嫩种子中表达量较高,说明pAhDGAT2a具有一定的组成型启动子活性。

关键词: 花生, DGAT2a基因, 启动子, 功能验证

Abstract:

Diacylglycerol acyltransferase (DGAT) is a rate-limiting enzyme in triacylglycerol (TAG) biosynthesis pathway. In this study, GenomeWalking method was used for cloning the promoter sequence of AhDGAT2a gene from Luhua 14, and finally a 1200 bp fragment flanking 5′-upstream of AhDGAT2a was obtained and named as pAhDGAT2a. The crucial regulatory elements in pAhDGAT2a were further analyzed with software PlantCARE. There were many TATA-box, CAAT-box, light regulation, stress and defense response and hormone response elements. To assess the activity of pAhDGAT2a, we constructed pAhDGAT2a:GUS cassettes and introduced it into the tobacco SR1 genome by Agrobacterium-mediated transformation. Expression pattern was monitored by histochemical staining. Results showed that GUS activity driven by the pAhDGAT2a was detected in almost all vegetative and reproductive tissues, with a higher expression level in stigma, anther and young seeds than in the other organs, indicating thatpAhDGAT2a has a constitutive promoter activity.

Key words: Arachis hypogaea L., DGAT2a gene, Promoter, Function analysis

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