【摘要】 参考家蝇防御素抗菌肽基因序列(AY260152)设计4条引物,用降落PCR方法获得家蝇防御素抗菌肽成熟肽基因。将基因克隆入酵母表达载体pPICZα-A,构建分泌型重组酵母表达载体pPIC-Des-HF,转化Pichia pastoris受体菌SMD1168,在醇氧化酶(AOX)启动子调控下,相对分子质量约5 000的重组抗菌肽Des-HF获得表达。抗菌试验结果表明,该表达产物对金黄色葡萄球菌有较好的抑菌活性。以小鼠为试验模型研究重组抗菌肽对金黄色葡萄球菌的体内抗菌活性,试验结果表明,240μg重组Des-HF可分别保护小鼠免受10倍最小致死浓度的金黄色葡萄球菌(ATCC26003)和CowanⅠ的攻击。更多还原

【Abstract】 With four pairs of specific primers designed according to the relevant sequence published on GenBank,the housefly antibacterial peptide defense gene was amplified with touch-down PCR method.After sequencing,the target gene was directly cloned into multiple cloning site of the pastoris expression plasmid pPICZα-A to get the recombinant plasmid pPIC-Des-HF.The 5 000 relative molecular weight recombinant antibacterial peptide Des-HF was secrectively expressed in the Pichia pastoris host cell SMD1168,when induced with methanol.The recombinant Des-HF showed perfect bacteriostasis activity against Gram-positive bacteria.Mice as animal model were used to test bacteriostasis activity and the result indicated that 240 μg recombinant Des-HF could completely protect the mice from ten folds minimum lethal dose concentration of Staphylococcus aureus(ATCC26003)and Cowan Ⅰ.更多还原