大豆蛋白磷酸酶基因GmPP2C28的克隆与表达分析

Cloning and Expression Analysis of the Protein Phosphatase Gene GmPP2C28 of Soybean

为解析植物蛋白磷酸酶2C(PP2C)在豆科植物与根瘤菌共生互作过程中的作用,以大豆为研究材料,克隆了大豆的一个蛋白磷酸酶基因GmPP2C28,并对其编码蛋白进行了亚细胞定位,随后在大肠杆菌中对其重组蛋白进行了表达和纯化。此外,通过荧光定量PCR技术,分析了GmPP2C28基因在大豆不同组织及接种根瘤菌后不同时期根系中的表达模式。结果表明,GmPP2C28完整编码区的cDNA序列长度为1029 bp,编码343个氨基酸,其编码蛋白定位于拟南芥原生质体的细胞核。体外重组蛋白以可溶形式高效表达,目的条带大小为37 kD左右。GmPP2C28基因在大豆的根和根瘤中表达水平较高,在接种根瘤菌后的大豆根中表达水平呈现先升高再下降的表达模式。

To elucidate the role of plant protein phosphatase 2C(PP2C)in the symbiotic interaction between leguminous plants and rhizobia,a protein phosphatase gene GmPP2C28 from soybean was cloned and its encoded protein was subcellular localized.Subsequently,the recombinant protein was expressed and purified in Escherichia coli.In addition,the expression patterns of GmPP2C28 gene in different tissues and roots at different stages after inoculation with rhizobia were analyzed using fluorescence quantitative PCR technology.The results showed that the cDNA sequence length of the complete coding region of GmPP2C28 was 1029 bp,encoding 343 amino acids,and its coding protein was located in the nucleus of Arabidopsis protoplasts.The recombinant protein in vitro was highly expressed in soluble form.The target band size of the purified GmPP2C28 protein was about 37 kD.The expression level of GmPP2C28 gene was relatively high in the root and nodule of soybean,and the expression level showed a pattern of first increasing and then decreasing in the root of soybean after inoculation with rhizobia.