酿酒葡萄分支酸合成酶基因(VvCS)的克隆与表达分析

Cloning and Expression Analysis of Gene Encoding Chorismate Synthase in Winegrapes

本研究以酿酒葡萄(Vitis vinifera)品种赤霞珠(Cabernet Sauvignon)及霞多丽(Chardonnay)为试材,采用in silico克隆和分子克隆相结合的策略,从果实中克隆到分支酸合成酶基因,命名为VvCS。该基因的cDNA编码区全长1312bp,编码436个氨基酸残基,预测其编码蛋白质分子量为46.9kD,等电点为7.8;生物信息学分析显示VvCS的DNA全长7117bp,包含13个外显子和12个内含子,定位于葡萄的第13号染色体上。VvCS编码的蛋白与其它植物来源的分支酸合成酶在氨基酸水平上的同源性为75%左右;实时荧光定量PCR分析表明VvCS在葡萄果实、茎、叶和叶柄组织中均有表达,且在果皮、果肉和种子中的表达变化趋势相似,与盛花后5周的果实相比,盛花后11周果实各部位中VvCS表达丰度均有不同程度增加。

A gene encoding chorismate synthase,named VvCS,was cloned from winegrape berries （Vitis vinifera） through adopting both in silico cloning and molecular cloning strategies.The full-length cDNA of VvCS contained an open reading frame of 1 312 bp in length,which encoded a polypeptide of 436 amino acid residues with a molecular mass of 46.9 kD and a pI of 7.8.Bioinformatics analysis showed that the genomic VvCS gene from genomic DNA locating on the 13th chromosome consisted of 7 117 bp in length,including 13 exons and 12 introns.The sequence homology analysis revealed that the VvCS had the homology of approximately 75% at amino acid level comparing with other plant CS proteins deposited in GenBank.Real time PCR analysis exhibited the VvCS gene transcriptionally expressed in all the tested tissues including fruits,stems,leaves and petioles,and similar trends in the transcript abundance were observed amongst the skin,pulp and seeds.Furthermore,the transcript abundance of these three tissues in the berries of 11 weeks after fully flowering showed relatively higher than that of the berries of 5 weeks after fully flowering.