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应用BlastX检索丹参毛状根的EST数据库,发现1条与分支酸变位酶(chorismate mutase,CM)高度同源的序列,克隆得到其全长cDNA,命名为SmCM1,Genbank登录号为KC784342。SmCM1全长948 bp,理论pI 6.41,与矮牵牛Petunia×hybrida、拟南芥Arabidopsis thaliana和毛果杨Populus trichocarpa的CM1分别具有70%,72%,64%的相似性。实时荧光定量PCR(real time quantitative PCR,QRT-PCR)分析表明SmCM1在丹参叶中的含量较高,其次是茎,在根中的含量相对较低。酵母诱导子(YE)和离子(Ag+)联合诱导丹参毛状根后,SmCM1与莽草酸途径的关键酶3-脱氧-D-阿拉伯-庚酮糖-7-磷酸合酶(3-deoxy-7-phosphoheptulonate synthase,DAHPS)和分支酸合酶(chorismate synthase,CS)基因表达趋势呈协同变化。YE+Ag+处理后3个基因皆上调表达,8 h达到表达高峰,分别是对照的7.9,5.5,9.8倍,随后下调,36 h时下降至对照水平以下,说明诱导处理之后糖代谢中间产物经莽草酸和分支酸途径合成苯丙氨酸和酪氨酸从而引起酚酸类化合物的过量积累。
BlastX was used to search the EST database of hairy root of Salvia miltiorrhiza and one highly homologous to chorismate mutase (CM) was found. The full-length cDNA was cloned and named as SmCM1. Genbank Accession No. KC784342. SmCM1 is 948 bp in length and has a theoretical pI of 6.41. It has 70%, 72% and 64% similarity to CM1 of petunia × hybrida, Arabidopsis thaliana and Populus trichocarpa, respectively. Real-time quantitative PCR (QRT-PCR) analysis showed that the content of SmCM1 in Salvia miltiorrhiza leaves was higher, followed by stems, and its content in roots was relatively lower. Induction of the hairy roots of Salvia miltiorrhiza by the combination of yeast elicitor (YE) and ion (Ag +) led to the interaction between SmCM1 and 3-deoxy-7, a key enzyme in the shikimic acid pathway -phosphoheptulonate synthase (DAHPS) and chorismate synthase (CS) genes showed synergistic changes. After YE + Ag + treatment, the three genes were up-regulated and peaked at 8 h, which were respectively 7.9, 5.5 and 9.8 times of the control, then down-regulated and below 36 h after the treatment, indicating that after induction treatment, the glucose metabolism intermediate product Shikimate and chorismate pathways synthesize phenylalanine and tyrosine to cause excessive accumulation of phenolic acids.