脱氧核酶抑制细菌TEM-1 β-内酰胺酶基因表达的实验研究  

Inhibitory effects of deoxyribozyme on bla_(TEM-1) gene expression in E.coli

作　　者：刘刚[1] 凌保东[1] 谢勇恩[1] 林丽[1] 赵廷坤[1] 雷军[1] 

机构地区：[1]川北医学院药物研究所,南充637007

出　　处：《中国抗生素杂志》2007年第11期693-696,共4页Chinese Journal of Antibiotics

基　　金：四川省教育厅自然科学科研项目(No2004A071);四川省重点科技项目(2006J13-080)

摘　　要：目的观察10-23脱氧核酶对细菌TEM-1酶基因表达的抑制作用。方法PCR扩增TEM-1酶全编码基因,将其连接入pBK-CMV载体中,并在大肠埃希菌JM109中表达。设计并合成针对blaTEM-1的10-23脱氧核酶、反义寡核苷酸,采用氯化钙法将其分别导入表达blaTEM-1的大肠埃希菌中,观察细菌导入脱氧核酶后,在含氨苄西林(100μg/ml)培养基中的生长活力及β-内酰胺酶活性的变化。结果10-23脱氧核酶导入表达blaTEM-1的大肠埃希菌后,在含氨苄西林(100μg/ml)的培养基中大肠埃希菌的生长活力明显低于反义寡核苷酸和空白对照组,导入脱氧核酶的大肠埃希菌β-内酰胺酶活性也明显低于反义寡核苷酸和空白对照组。结论10-23脱氧核酶能特异性地抑制细菌blaTEM-1酶基因表达。Objective To investigate blaTEM-1 gene expression in E. coli. Methods the inhibitory effects of 10-23 deoxyribozyme （10-23 DRz） on blaTEM-1 gene was amplified by polymerase chain reaction （PCR）, the target amplification fragment was cloned into PBK-CMV vector. The recombinant vector was sequenced by Sanger＇s dideoxy chain termination composition method. According to the gene sequence of blaTEM-1, 10-23 DRz and antisense oligonucleotides （As-ODN） were designed, synthesized, and introduced into E. coli producing TEM-1 respectively by the CaCl2 procedure, bacterial viability in liquid medium containing ampicillin （100 μg/ml） and β-lactamase activity were detected spectrophotometrically. Results A600 value of E. coli incubated with 10-23 DRz was lower, compared with that of corresponding group （P〈0. 05）. β-lactamase activity of 10- 23 DRz group was also lower than that of corresponding group （P〈0. 01）. Conclusion 10-23 DRz could specifically inhibit the blaTEM-1 expression in E. coll.

关 键 词：耐药性 Β-内酰胺酶 脱氧核酶 基因表达 

分 类 号：R978.11[医药卫生—药品]