McDonald G A, Anacker R L, Garjian K
Science. 1987 Jan 2;235(4784):83-5. doi: 10.1126/science.3099387.
Two major protective antigens of Rickettsia rickettsii have been previously described. In this study, we cloned the gene encoding one of these antigens into Escherichia coli and tested the effectiveness of the recombinant-made product as a vaccine for Rocky Mountain spotted fever. A clone bank of R strain R. rickettsii DNA was made in E. coli K-12 by using the plasmid vector pBR322. Transformants were screened for their ability to make rickettsial antigens by reactivity with rabbit antibodies to R. rickettsii. One of the transformants, EM24(pGAM21), made a product reactive with two monoclonal antibodies that recognize a 155-kilodalton protein of R. rickettsii. One of the monoclonal antibodies was a member of a class of antibodies that react to heat-sensitive epitopes and protect mice injected with a potentially lethal dose of viable R. rickettsii. The cloned product contained this protective heat-sensitive epitope. In order to obtain enhanced expression, the gene was subcloned downstream of the lactose promoter on the plasmid vector pUC8. A sonic lysate of E. coli harboring the pUC8 subclone was used successfully as a vaccine to protect mice injected with a lethal dose of the viable R. rickettsii.
此前已描述了立氏立克次体的两种主要保护性抗原。在本研究中，我们将编码其中一种抗原的基因克隆到大肠杆菌中，并测试了重组制成的产品作为落基山斑疹热疫苗的有效性。通过使用质粒载体pBR322，在大肠杆菌K-12中构建了立氏立克次体R株DNA的克隆文库。通过与抗立氏立克次体的兔抗体反应，筛选转化体产生立克次体抗原的能力。其中一个转化体EM24(pGAM21)产生了一种与两种单克隆抗体反应的产物，这两种单克隆抗体识别立氏立克次体的一种155千道尔顿的蛋白质。其中一种单克隆抗体是一类与热敏感表位反应的抗体成员，可保护注射了潜在致死剂量活立氏立克次体的小鼠。克隆产物包含这种保护性热敏感表位。为了获得增强表达，该基因被亚克隆到质粒载体pUC8上乳糖启动子的下游。携带pUC8亚克隆的大肠杆菌的超声裂解物成功用作疫苗，以保护注射了致死剂量活立氏立克次体的小鼠。
