大豆候选草酸氧化酶基因片段克隆与表达分析

The Cloning and Expression Analysis of Putative Oxalate Oxidase Gene in Soybean

为克隆大豆草酸氧化酶基因并初步鉴定其功能,根据双子叶植物的草酸氧化酶基因,设计简并引物,同源克隆大豆该基因片段,实时定量PCR分析基因表达情况。序列分析发现,基因片段大小504bp,无内含子,编码168个氨基酸;Blastx同源比对发现,与大豆的germin基因和苜蓿、花生的草酸氧化酶基因高度同源;草酸诱导大豆耐病和感病种质叶片基因表达分析表明,基因的表达与草酸诱导有关,而且在耐菌核病种质与感菌核病种质间,表达量和表达时间差异明显。推断该基因片段可能是一具有草酸氧化酶活性的大豆germin基因片段。

In order to clone oxalate oxidase gene of soybean and identify its function, the degenerate primer was designed on based on homology oxalate oxidase genes of dicotyledonous plant from GenBank database. Real-time quantitative PCR was used to analyze the gene expression. The cloned gene fragment size was 504 bp without intron and encoded 168 amino acids. Homology comparison found it was high homology with germin protein of soybean and oxalate oxidase of medicago and arachis hypogaca. The gene expression induced by oxalate showed that the expression was relative with oxalate induction. There was significant difference in expression amount and expression time between resistant and susceptive germplasm. To infer that the gene fragment was a germin gene fragment with oxalate oxidase activity.