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期 刊 ：植物遗传资源学报 2014年 15卷 2期

关键词：MdKS；MdKOA1；克隆；亚细胞定位；表达；

Keywords:MdKS, MdKOA1, cloning, subcellular location, expression,

摘 要 ：内根-贝壳杉烯合成酶基因（KS）、内根-贝壳杉烯酸氧化酶基因（KOA）是赤霉素合成途径的关键基因。通过touchdown PCR从‘短枝富士’茎尖组织中克隆得到KS和KOA基因的开放阅读框（ORF），分别命名为MdKS（GenBank登录号KF437681）和MdKOA1（GenBank登录号为KF437682）。MdKS ORF长2211bp，MdKOA1 ORF长1509bp。氨基酸同源性分析表明，MdKS与已报道的其他植物物种的氨基酸序列具有45.2%-98.8%的同源性；MdKOA1与其他物种的氨基酸序列同源性为42.8%-99.4%。洋葱表皮的瞬时表达显示，MdKS蛋白定位于细胞核和细胞质；MdKOA1蛋白定位于细胞质中。荧光定量结果表明，MdKS与MdKOA1基因在SH40、SH28嫁接品种‘短枝富士’空间组织中的表达模式基本是一致的。在茎尖、幼果、枝皮中半矮化类型SH28嫁接‘短枝富士 ’MdKS和MdKOA1基因的表达量高于矮化类型SH40嫁接富士。

Abstract:Ent-kaurene Synthase (KS) and ent-kaurenoic acid oxidase gene (KOA) are critical genes in the pathway of gibberellins biosynthesis. In this research, KS and KOA gene in apple, designated as MdKS (GenBank accession number: KC437681）and MdKOA1 (GenBank accession number: KC437682), were isolated from the apical tissue of ‘spur-type Fuji’ apple by touchdown PCR. The length of MdKS ORF is 2211bp, MdKOA1 ORF is 1509bp. Amino acid sequence analysis revealed that the KS sequence had 45.2%-98.8% similarity with those of other report plant; and the KOA sequence had 42.8-99.4% similarity with other plant. Subcelluar localization assays that the MdKS protein was located in the nucleus and the plasma; and the MdKOA1 protein located in the plasma. Fluorescent quantitative PCR analysis showed that the trend of expression of MdKS and MdKOA1 gene are the same in SH40 and SH28 grafted cultivar ‘spur-type Fuji’. The expression of MdKS and MdKOA1 gene in the apical tissue, young fruit and branch bark of semi-dwarf type SH28 grafted cultivar ‘spur-type Fuji’ are higher than dwarf type SH40 grafted cultivar .

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