农业生物技术学报2011，Vol.19Issue(2)：258-262,5.DOI:10.3969/j.issn.1674-7968.2011.02.009
番茄Metacaspase基因(LeM)的克隆及其在大肠杆菌Rosetta中的高效表达
Cloning of Lycopersicon esculentum Metacaspase Gene (LeM) and Its Effective Expression in Escherichia coli Rosetta
摘要
Abstract
To obtain the fusion protein of Lyco persicon esculentum Metacaspase (LeM) and study the characteristics and function of it, we cloned the open reading frame of LeM in Lycopersicon esculentum by RT-PCR, and successfully constructed the prokaryotic expression vector pET28a-LeM for LeM expression.The recombinant plasmid pET28a-LeM was transformed into Escherichia coli BL21 (DE3)PlysS and Rosette for expression, induced by IPTG, and the conditions of the expression was optimized.SDS-PAGE revealed that the best expression was induced by 1 mmol/L IPTG at 37℃ and the recombinant protein expressed mainly as inclusion body in E.coli BL21(DE3)PlysS while as fusion protein in E.coli Rosetta at the same condition.Enzyme activity assay showed that the activity of LeM in E.coli Rosetta was 33 RFU/min, which was much higher than 7 RFU/min of E.coli BL21 (DE3)PlysS.We demonstrate that the strain has a great influence on the expression of LeM fusion protein.关键词
番茄/Metacaspase/基因克隆/原核表达Key words
Lycopersicon.esculentum/ Metacaspase/ Gene cloning/ Prokaryotic expression引用本文复制引用
马秋敏,张雅丽,刘金娜,车英慧,谢寒丁,罗云波,曲桂芹..番茄Metacaspase基因(LeM)的克隆及其在大肠杆菌Rosetta中的高效表达[J].农业生物技术学报,2011,19(2):258-262,5.基金项目
本研究由中央高校基本科研业务费专项资金(No.2009-1-83)、国家公益性行业(农业)科研专项项(No.200803033-B0602-03)和国家自然科学基金(No.30500352)共同资助 （No.2009-1-83）
