Bagdasarian M M, Amann E, Lurz R, Rückert B, Bagdasarian M
Gene. 1983 Dec;26(2-3):273-82. doi: 10.1016/0378-1119(83)90197-x.
A broad-host-range vector, pKT240, containing the structural gene (aph) for aminoglycoside phosphotransferase (APH), without promoter, has been constructed. Insertion of DNA fragments carrying promoters upstream of aph gene into the unique EcoRI site of this vector results in the expression of the aph gene and consequently the resistance of the host cells to streptomycin. The new vector has been used to show that the hybrid trp-lac (tac) promoter and the promoter of the lacIQ gene of Escherichia coli are active in Pseudomonas putida. Derivatives of pKT240 containing tac and lacIQ sequences may be used as wide-host-range expression vectors. Regulated overproduction of APH and catechol 2,3-oxygenase can be obtained with the aid of the new vectors in both E. coli and P. putida.
构建了一种广宿主范围载体pKT240，其含有氨基糖苷磷酸转移酶（APH）的结构基因（aph），但无启动子。将携带启动子的DNA片段插入aph基因上游的该载体独特的EcoRI位点，导致aph基因表达，从而使宿主细胞对链霉素产生抗性。该新型载体已用于证明杂交trp-lac（tac）启动子和大肠杆菌lacIQ基因的启动子在恶臭假单胞菌中具有活性。含有tac和lacIQ序列的pKT240衍生物可用作广宿主范围表达载体。借助新型载体，在大肠杆菌和恶臭假单胞菌中均可实现APH和儿茶酚2,3-双加氧酶的调控过量表达。
