Cloning and Expression Analysis of Gene Encoding Shikimate Kinase in Grapes
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摘要: 以"赤霞珠"葡萄(Vitis vinifera)果实为实验材料,采用电子克隆和分子克隆相结合的方法,克隆到莽草酸激酶基因,命名为VvSK。VvSK的cDNA编码区全长906bp,编码301个氨基酸残基。预测该蛋白质相对分子质量为33.2×103,等电点为8.6,VvSK的DNA全长4309bp,包含10个外显子和9个内含子,定位于7号染色体上。VvSK编码的蛋白与其他植物中同类酶在氨基酸水平上的同源性最高达64.52%。系统进化树分析显示VvSK与毛果杨SK基因亲缘关系较近。荧光实时定量PCR分析结果表明,VvSK在葡萄果实、茎、叶和叶柄组织中均有表达,在不同发育期果实的果皮、果肉和种子中相对表达量存在差异,与UV-A和UV-B照射处理相比,UV-C照射对VvSK基因表达的诱导效应较明显。Abstract: A shikimate kinase gene named VvSK was cloned from grape（Vitis vinifera L.Cabernet Sauvignon） berries by means of both in silico cloning and molecular cloning.The full-length cDNA of VvSK contains an open reading frame of 906 bp which encodes a polypeptide of 301 amino acid residues with a molecular mass of 33.2 kD and a pI value of 8.6.The VvSK gene from genomic DNA located on the 7th chromosome consists of 4 309 bp,with 10 exons and 9 introns.The sequence homology comparison revealed that the VvSK sequence exhibits a high homology（upto 64.52%） at amino acid level with other plant SK proteins from the GenBank.Phylogenetic analysis indicated that the VvSK has a close evolutionary relationship with Populus trichocarpa SK.The analysis of real-time PCR showed the transcriptional expression of VvSK was detected in all the tested tissues including fruit,stem,leaf and petiole.But some differences in transcript abundance were detected in the skin,pulp and seeds at the different fruit development stages.UV-C treatment induced higher expression of VvSK in the 3-week and the 11-week berries than other treatments（UV-A and UV-B）.
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Key words:
- shikimate kinase（SK） /
- cloning /
- gene expression /
- Vitis vinifera
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