摘要
为了获得纯化的长喙田菁(Sesbania rostrata)植物螯合肽合成酶PCS2,以原核表达载体pMAL-c2x为基础,构建了含有SrPCS2开放阅读框序列的原核表达载体pAM57,将其转化表达菌株BL21(DE3),对融合蛋白的表达进行了优化,通过Western blotting鉴定融合蛋白,并用麦芽糖亲和柱对MBP-SrPCS2融合蛋白进行纯化.结果表明:在15℃下,经0.2mmol/L IPTG诱导16h可以表达出可溶性的MBP-SrPCS2融合蛋白,通过亲和层析得到了纯化的MBP-SrPCS2融合蛋白,为进一步研究SrPCS2的活性及PCS的催化机制奠定了基础.
The mature peptide of Sesbania rostrata phytochelatin synthase2（SrPCS2） was subcloned into pMAL-c2x vector to construct the prokaryotic expression vector pAM57.The recombinant plasmid was transformed into E.coli BL21（DE3）,then the fused protein MBP-SrPCS2 was induced by IPTG at different temperature,identified by Western blotting and purified through Maltose Binding column.The results suggested that the soluble fusion protein MBP-SrPCS2 was expressed at high levels 16 h after 0.2 mmol/L of IPTG induction（15℃） and the purified MBP-SrPCS2 was obtained by affinity chromatography.
出处
《甘肃农业大学学报》
CAS
CSCD
北大核心
2011年第6期150-154,共5页
Journal of Gansu Agricultural University
基金
湖北省重点(培育)学科:农业资源利用(0903)
湖北省自然科学基金项目(2005ABA083)
