【摘要】 Fe-S簇可参与电子传递和基因调节等过程.实验以嗜热菌Thermus thermophilus HB8基因组为模板,通过PCR扩增获得Fe-S簇ISC系统生物合成途径中的IscA和IscU基因,并克隆连接到表达载体pET-28a(+)上.重组质粒转化E.coli Rosetta(DE3)进行表达,用亲和层析纯化得到浓度和纯度都较高的IscA和IscU蛋白.SDS-PAGE分析,证实其分子量大小分别为13.7KD和14.8KD.实验确定IscA是单聚体,IscA上能形成铁硫簇,IscU上不能形成铁硫簇.在厌氧条件下,RNA的量能提高1.5倍,但是IscA和IscU的表达量都会降低,说明厌氧对这两个基因有损伤.更多还原

【Abstract】 Iron-sulfur cluster plays an important role in electron transfer and gene regulation. In this study,the gene of IscA and IscU from HB8 were cloned and expressed in Rosetta,and the proteins were purified by using affinity chromatography. SDS-PAGE revealed the moleculer weight of IscA was 13. 7KD and IscU was 14. 8KD. Iron sulfur cluster can be formed on IscA,Iron sulfur cluster can’t be formed on IscU,under anaerobic conditions,RNA can increase 1. 5 times,But the amount of expression of IscA and IscU will be reduced,indicating that the anaerobic damage on these two gene.更多还原