Wang Zhengjun, Ge Qian, Chen Chen, Jin Xinxin, Cao Xiaoyan, Wang Zhezhi
Key Laboratory of the Ministry of Education for Medicinal Resources and Natural Pharmaceutical Chemistry, National Engineering Laboratory for Resource Development of Endangered Crude Drugs in Northwest of China, College of Life Sciences, Shaanxi Normal University, Xi'an, 710119, People's Republic of China.
The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an, 710049, China.
Appl Biochem Biotechnol. 2017 Feb;181(2):562-572. doi: 10.1007/s12010-016-2231-4. Epub 2016 Sep 9.
In this study, we cloned a full-length cDNA and the genomic DNA sequence of SmCCoAOMT (GenBank ID JQ007585) from Salvia miltiorrhiza. The 744-bp open-reading frame encodes a protein of 247 amino acids that shares 95 % similarity with one in Vitis vinifera. Real-time quantitative PCR analysis revealed that SmCCoAOMT is most highly expressed in the stems and can be induced by methyl jasmonate (MeJA) and XC-1 treatment. To evaluate its function in vivo, we generated RNA interference transgenic plants through Agrobacterium tumefaciens-mediated gene transfer. Compared with untransformed control plants, the transgenics had significantly less lignin and the expression of lignin-biosynthetic genes SmCCR and SmCOMT was depressed. In 90-day-old roots from plants of transgenic line M5, accumulations of rosmarinic acid and salvianolic acid B (Sal B) were greatly reduced by 0.89- and 0.69-fold, respectively. This low-Sal B phenotype was stable in the roots, with the level of accumulation being approximately 43.58 mg g dry weight, which was 52 % of the amount measured in the untransformed control. Our results suggest that SmCCoAOMT is involved in lignin biosynthesis and affects the accumulation of phenolic acids. This study also provides potential guidance for using lignin-related genes to genetically engineer Salvia miltiorrhiza.
在本研究中,我们从丹参中克隆了一个全长cDNA以及SmCCoAOMT(GenBank ID JQ007585)的基因组DNA序列。744碱基对的开放阅读框编码一个由247个氨基酸组成的蛋白质,该蛋白质与葡萄中的一种蛋白质具有95%的相似性。实时定量PCR分析表明,SmCCoAOMT在茎中表达量最高,并且可被茉莉酸甲酯(MeJA)和XC-1处理诱导。为了评估其在体内的功能,我们通过根癌农杆菌介导的基因转移产生了RNA干扰转基因植株。与未转化的对照植株相比,转基因植株的木质素显著减少,木质素生物合成基因SmCCR和SmCOMT的表达受到抑制。在转基因株系M5植株90日龄的根中,迷迭香酸和丹酚酸B(Sal B)的积累量分别大幅降低了0.89倍和0.69倍。这种低Sal B表型在根中是稳定的,积累水平约为43.58 mg g干重,是未转化对照中测量量的52%。我们的结果表明,SmCCoAOMT参与木质素生物合成并影响酚酸的积累。本研究还为利用木质素相关基因对丹参进行基因工程改造提供了潜在指导。