Chatterji U, Ahmad R, Venkaiah B, Hasnain S E
Eukaryotic Gene Expression Laboratory, National Institute of Immunology, New Delhi, India.
Gene. 1996 Jun 1;171(2):209-13. doi: 10.1016/0378-1119(96)00188-6.
The baculovirus system is an extremely powerful tool for expression of heterologous genes in eukaryotic environment. A multiple expression vector, pBacUCmP3, was constructed which harbored two copies of the Autographa californica nuclear polyhedrosis virus very late gene promoter and the Drosophila melanogaster 70-kDa heat-shock protein (hsp70) promoter with downstream unique restriction sites for cloning of three independent foreign genes. Co-transfection of pBacUCmP3 with Bsu36I-linearized viral DNA yields recombinant progeny viruses at very high frequencies. The utility of this multiple expression transfer vector was demonstrated using three heterologous reporter genes encoding the beta-subunit of the human chorionic gonadotropin hormone, firefly luciferase and the bacterial beta-galactosidase (beta Gal) enzyme. The expression of reporter genes, monitored at various times post-infection, confirmed that while beta-Gal synthesis was under the transcriptional control of the hsp70 promoter, the beta hCG and Luc proteins were synthesized as a function of polyhedrin promoter activation profile. This vector will be useful for multiple synthesis of proteins at different time points.
杆状病毒系统是在真核环境中表达异源基因的一种极其强大的工具。构建了一个多表达载体pBacUCmP3，它含有两份苜蓿银纹夜蛾核型多角体病毒极晚期基因启动子以及果蝇70 kDa热休克蛋白（hsp70）启动子，并在下游带有用于克隆三个独立外源基因的独特限制酶切位点。用Bsu36I线性化的病毒DNA与pBacUCmP3共转染能以非常高的频率产生重组子代病毒。使用编码人绒毛膜促性腺激素β亚基、萤火虫荧光素酶和细菌β-半乳糖苷酶（β Gal）的三个异源报告基因证明了这种多表达转移载体的实用性。在感染后的不同时间监测报告基因的表达，证实虽然β-Gal的合成受hsp70启动子的转录控制，但β hCG和Luc蛋白的合成是多角体蛋白启动子激活情况的函数。该载体将有助于在不同时间点进行多种蛋白质的合成。
