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青花菜β-1,3-葡聚糖酶基因的克隆、表达分析及其植物表达载体的构建

DOI: 10.7685/j.issn.1000-2030.2008.02.006, PP. 27-31

Keywords: 青花菜,霜霉病,β-1,3-葡聚糖酶基因,植物表达载体

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Abstract:

根据其他植物的β-1,3-葡聚糖酶基因保守域序列设计简并引物,扩增出青花菜β-1,3-葡聚糖酶基因的中心区段,结合5′RACE和3′RACE技术获得该基因的5′端序列和3′端序列,经序列拼接获得1个1277bp的cDNA,在GenBank中登录号为EF484879,定名为BObg。序列分析结果表明,该cDNA包含1个1056bp的开放阅读框,编码352个氨基酸,其理论上的等电点PI=7.314,相对分子质量为3.892×104,聚类分析显示该序列与已报道的其他植物的β-1,3-葡聚糖酶基因具有较高氨基酸序列同源性。利用未接种和接种霜霉病后不同时期的青花菜植株细胞进行半定量RT-PCR分析,结果表明,该基因在霜霉病接种后48h优势转录表达,未接种和接种后其他时期都呈低水平转录表达。利用pBI121质粒构建含35S启动子的正义表达载体,酶切图谱和PCR分析结果证明该表达载体构建成功。

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