小麦ent-柯巴基焦磷酸合酶基因TaCPS1的克隆与荧光定量分析

Cloning and qRT-PCR Analysis of TaCPS1 Gene in Wheat

ent-柯巴基焦磷酸合酶(ent-copalyl diphosphate synthase,CPS)是植保素合成途径中的关键酶。本研究利用mRNA差异显示技术,从小偃麦异附加系种质SN6306中得到一个与抗白粉病相关的Ta CPS1基因。该基因ORF长2 394 bp,编码797个氨基酸。对其进行生物信息学分析表明,该序列推导的蛋白无信号肽,主要由亲水性氨基酸组成,可能存在于细胞质中;其具有类异戊二烯合成酶超家族的保守结构域。荧光定量分析结果表明,SN6306中的Ta CPS1基因在白粉病菌E09诱导处理72 h时,表达量上调了大约45倍,但感病亲本烟农15(YN15)基本不受诱导。在茉莉酸甲酯诱导下,Ta CPS1基因表达量升高将近15倍;在水杨酸诱导下,Ta CPS1基因表达量升高将近2.4倍。推测Ta CPS1基因可能参与小麦抗白粉病机制中的水杨酸和茉莉酸激素调节途径。

ent- copalyl diphosphate synthase（ CPS） is a key enzyme in the biosynthesis of phytoalexins. In this study,the c DNA of Ta CPS1 gene which was related to resistance to powdery mildew was identified from the wheat- Thinopyrum alien addition line SN6306 by mRNA differential display. The c DNA had the length of 2 394 bp,which encoded 797 amino acid residues. Bioinformatics analysis indicated that the encoded protein which was mainly consisted of hydrophilic amino acids did not contain signal peptide. It was located in cytoplasm belonging to the Isoprenoid- Biosyn- C1 superfamily. qRT- PCR analysis showed that the expression of Ta CPS1 gene in SN6306 was up- regulated about 45 times after induced by E09 for 72 hours,while that in YN15 was basically not induced. The expression of Ta CPS1 increased by nearly 15 times and2. 4 times under the induction of JAMe and SA respectively. Thus we hypothesized that Ta CPS1 might be involved in the regulation pathway of SA and JA in the powdery mildew resistance mechanism.