Tang Yaping, Yuan Hui, Qin Jianbing
Research Group of Molecular Biology and Informatics, Xinjiang Normal University, Urumqi 830053, Xinjiang, China.
Sheng Wu Gong Cheng Xue Bao. 2012 Jun;28(6):705-14.
The flavonoid-3-O-glucosyltransferas (3GT) is one of the most important enzymes for biosynthesis of plant secondary metabolites. In this paper, the homology analysis and gene special primers design were used. With the methods of modern molecular biology, the full-length gene of 3GT (GenBank Accession No. JN092127) which was cloned from Saussurea involucrata Kar. et Kir by RT-PCR and RACE. The cDNA sequence of 3GT consisted of 1548 bp open reading frame (ORF) encoding 516 amino acid, the deduced 3GT protein shared 91% and 89% identities with that of Fragaria x ananassa GT6, Manihot esculenta anthocyanidin 3-O-glucosyltransferase. Homology analysis showed that deduced 3GT protein has a glycosyltransferase signature domain PSPG-box. The transcripts of 3GT members were found mainly in leaves and callus. 3GT gene of S. involucrata was under the control of the cauliflower mosaic virus (CaMV) 35S promoter, homologous transformation used an Agrobacterium rihizogenes-mediated transformation system. The results on UV spectrophotometry showed S. involucrata callus after suspension culture that an average of total flavonoids on transgenic callus was 2.06 times higher than non-transgenic callus.
类黄酮 -3-O-葡萄糖基转移酶(3GT)是植物次生代谢产物生物合成中最重要的酶之一。本文采用同源性分析和基因特异性引物设计方法。运用现代分子生物学手段,通过RT-PCR和RACE技术从新疆雪莲(Saussurea involucrata Kar. et Kir)中克隆得到3GT全长基因(GenBank登录号:JN092127)。3GT的cDNA序列包含1548 bp的开放阅读框(ORF),编码516个氨基酸,推导的3GT蛋白与凤梨草莓GT6、木薯花青素3-O-葡萄糖基转移酶的一致性分别为91%和89%。同源性分析表明,推导的3GT蛋白具有糖基转移酶特征结构域PSPG-box。3GT成员的转录本主要在叶片和愈伤组织中被发现。新疆雪莲3GT基因受花椰菜花叶病毒(CaMV)35S启动子控制,采用发根农杆菌介导的转化系统进行同源转化。紫外分光光度法检测结果显示,悬浮培养后的新疆雪莲愈伤组织中,转基因愈伤组织总黄酮平均含量比非转基因愈伤组织高2.06倍。