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大豆科学  2014 

大豆GmSOT1基因克隆、载体构建及初步表达研究

DOI: 10.11861/j.issn.1000-9841.2014.06.0815, PP. 815-819

Keywords: 大豆,磺酸基转移酶,硫代葡萄糖苷,基因克隆,载体构建

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Abstract:

磺酸基转移酶(sulphotransferase,简称SOT)是硫代葡萄糖苷(glucosinolate,简称硫苷)核心结构合成途径中的关键酶之一。本研究采用反向遗传学方法从大豆基因组中克隆GmSOT1基因并对其功能进行初步研究。首先采用PCR方法扩增大豆-GmSOT1-基因,并将其连接到pRNAi载体上的35S(CaMV35S)启动子后面;将35S:GmSOT1-基因片段从pRNAi载体上双酶切下来,连接到植物双元表达载体pCAMBIA2301的多克隆位点上,且命名为pCAMBIA2301-35S:GmSOT1;采用根癌农杆菌介导的遗传转化方法转化大豆成熟胚,并考察其初步表达结果。结果表明:大豆-GmSOT1-基因的编码区(codingsequences,简称CDS)由1014bp的核苷酸组成,编码一个长度为337个氨基酸残基的蛋白质;35S:GmSOT1被成功导入大豆胚细胞,初步观察发现GmSOT1过量表达导致大豆胚根明显膨大弯曲。

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