葡萄果实分支酸变位酶基因的克隆与表达分析

Cloning and expression of gene encoding chorismate mutase in grape berries

莽草酸途径是连接糖代谢和次生代谢的主要桥梁,分支酸变位酶(Chorismate Mutase,CM)是控制碳同化物由莽草酸途径进入苯丙烷代谢途径的入口酶,在葡萄果实酚类物质积累中起着重要作用。本研究以赤霞珠葡萄(Vitis vinifera cv.Cabernet Sauvignon)果实为试材,采用电子克隆和分子克隆相结合技术,获得两个编码分支酸变位酶同源基因,分别命名为VvCM1和VvCM2。这两个基因分别定位于4号和14号染色体上,其编码区全长分别为741 bp和963bp,编码蛋白含246和320个氨基酸。VvCM1和VvCM2与其他植物中的同类酶在氨基酸水平上具有广泛的同源性。Real-time PCR分析显示,VvCM1和VvCM2基因在葡萄的各器官和组织中均有表达,VvCM1在果实中表达丰度最高,而VvCM2在茎中表达丰度最高。

Shikimate pathway connects primary carbohydrate metabolism with the biosynthesis of most secondary metabolites in plants. Chorismate Mutase （CM） is an entry enzyme from＆amp;nbsp;shikimate pathway into phenylpropanoid metabolism, which plays an important role in the accumulation of phenolic compounds in grape berries. In the present study, two CM cDNAs were cloned from grape berries （Vitis vinifera cv. Cabernet Sauvignon） and designated as VvCM1 and VvCM2. The cDNAs of VvCM1 and VvCM2 contained open reading frames of 741 bp and 963 bp, which encoded a polypeptide of 246 and 320 amino acid residues with a molecular mass of 27.06 kDa and 35.2 kDa, respectively. The DNA sequences corresponding to the two isogenes both contain four introns and located on the chromosome 4 and 14, respectively. The sequence homology comparison showed that VvCM1 and VvCM2 had high homology of the amino acid sequences with other plant CM from the GenBank. Analysis by Real time-PCR showed that VvCM1 and VvCM2 were expressed in all the tested tissues. The transcript abundance of VvCM1 was the highest in grape berries and VvCM2 had the highest transcript level in stems.