Position: Home > Articles > Construction of E.coli-App temperature controlled double gene cracking vect or pMC-WK
Chinese Journal of Veterinary Science
2011,31
(9)
1304-1308
利用E.coli-App穿梭载体构建温控双基因裂解载体pMC-WK
作 者：
孙颖;金天明;冯立文;袁洁
单 位：
广东省农业科学院兽医研究所
关键词：
裂解蛋白E;核酸酶A;App菌影;穿梭载体
摘 要：
采用PCR方法从噬菌体PhiX174基因组中扩增出裂解蛋白E基因,从金黄色葡萄球菌基因组中扩增出核酸酶A基因(SN),通过15个柔性氨基酸linker将双基因串联,插入PBV220表达载体,构建温控双基因裂解系统(DLS)。PCR扩增DLS,将其与E.coli-App穿梭载体pMC-Express相连,构建重组穿梭载体pMC-WK。裂解试验表明,E-15aalinker-SN裂解较E裂解迅速而彻底,菌体裂解率达到99.999 4%;经PCR扩增出的DLS具有裂解活性。本试验成功构建了温控双基因裂解载体pMC-WK,为App菌影疫苗的研究奠定了基础。
译 名：
Construction of E.coli-App temperature controlled double gene cracking vect or pMC-WK
作 者：
SUN Ying1,JIN Tian-ming1,FENG Li-wen2,YUAN Jie3 (1.Animal Science Department,Tianjin Agricultural College,Tianjin 300 384,China;2.College of Aminal Science and Veterinary Medicine,Jilin University Changchun 130062,China;3.Institute of Guangdong Veterinary Science,Guangzhou 510640,China)
关键词：
lysis protein E;nuclease A;App bacterial ghost;shuttle vector
摘 要：
Using the PCR method to clone lysis protein E gene from the genome of phage PhiX174 and nuclease A gene(SN) from the genome of Staphylococcus aureus. Link the two genes with 15 amino acid flexible linker to get double-gene tandem, then insert it into PBV220 plasmid.Amplifing DLS through PCR to construct pMC-W K on the basis of E.coli-App shuttle vector pMC-Express.Lysis experiments s how th at E-15aalinker-SN system has better cracking efficiency than E gene system.Th e lysis rate can be 99.999 4% and DLS gene amplified by PCR with the cleavage acti vity.The test successfully constructed temperature controlled double gene cracki ng vector pMC-WK,making foundation for the App bacterial ghost vaccine research.
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