大豆GmOLPa基因的蛋白序列分析及原核表达

Protein Sequence Analysis and Prokaryotic Expression of GmOLPa Gene from Soybean

以盐诱导后的大豆根为材料,提取总RNA,RT-PCR得到GmOLPa目的片段。对其蛋白序列分析表明,该蛋白属于GH64-TLP-SF同源超家族中的PR-5蛋白,第25-244位氨基酸是其保守结构域,与番茄中的PR-5亲缘关系最为接近。构建GmOLPa基因ORF的原核表达载体pET-28-GP,并对其在大肠杆菌BL21中的表达条件进行优化。SDS-PAGE证实重组质粒能够在大肠杆菌BL21中表达,目的蛋白分子量约为30 kD,最佳诱导时间为4 h,最佳IPTG诱导浓度为1.0 mmol/L。

In order to identify the function of GmOLPa gene from soybean, total RNA was extracted from the root of soybean induced by salt stress and GmOLPa gene fragment was obtained by RT-PCR. GmOLPa protein sequence was analyzed and the results showed that GmOLPa protein belonged to PR-5 of GH64-TLP-SF homologous superfamily and was the most closely genetic relationship with the PR-5 from tomato. The 25-244 amino acids in the GmOLPa protein sequence were the conservative domain. Prokaryotic expression vector pET-28-GP of GmOLPa gene was constructed and expressive conditions of pET-28-GP in E. coli BL21 were optimized. SDS-PAGE was assayed and showed that recombinant plasmid could be inducted and expressed in E. coli. The target protein molecular weight was approximately 30 kD, the optimal induction time of 4 h and the optimal IPTG induction concentration of 1.0 mmol/L.