摘要
利用分子克隆的方法构建夏侧金盏花虾青素合成相关酶基因Adkc的双T-DNA表达载体。首先设计基因Adkc、启动子E8和终止子Hsp的引物,选用植物双T-DNA表达载体pGreen-DT。其次将E8、Adkc和Hsp按顺序连接到载体上,构建Adkc双T-DNA、果实特异表达的表达载体。使用Q5超保真酶成功把Adkc和Hsp扩增到一起,经酶切和连接后将启动子E8、Adkc和Hsp成功插入植物双T-DNA表达载体pGreen-DT上。成功构建夏侧金盏花虾青素合成相关酶基因Adkc的双T-DNA表达载体。为后续研究中将该载体转入植物中表达,分析Adkc的功能及其参与的代谢途径、获得高效表达虾青素且不含抗生素基因的转基因植株做好了前期准备。
A double T-DNA expression vector of Adkc,a related gene of astaxanthin from Adonis aestivalis,has been constructed by molecular cloning.Firstly,the primers of gene Adkc,promoter E8 and terminator Hsp were designed,and the double T-DNA expression vector pGreen-DT was used as the vector.Secondly,E8,Adkc and Hsp were connected into the vector in order to construct the fruit-specific expression vector of Adkc with double T-DNA sequences.Adkc and Hsp were successfully amplified by using Q5 high-fidelity enzyme.After enzyme digestion and ligation,the promoter E8,Adkc and Hsp were successfully inserted into the plant double T-DNA expression vector p Green-DT.This study laid the foundation for the follow-up research of transferring the vector into a plant to analyze the function of Adkc and its metabolic pathway,and to obtain transgenic plants with high-efficiency expression of astaxanthin without antibiotic genes.
作者
黄静
常沙沙
杨卫丽
曾玉
高新征
Huang Jing;Chang Shasha;Yang Weili;Zeng Yu;Gao Xinzheng(School of Basic Medical Science,Hainan Medical University,Haikou,571199;Pharmaceutical College,Hainan Medical University,Haikou,571199)
出处
《分子植物育种》
CAS
CSCD
北大核心
2019年第22期7386-7389,共4页
Molecular Plant Breeding
基金
海南省自然科学基金面上项目(20163077)
海南省科协青年科技英才学术创新计划项目(HAST201633)共同资助
