Molecular cloning and expression of Treponema pallidum DNA in Escherichia coli K12.
A gene bank of Treponema pallidum DNA in Escherichia coli K-12 was constructed by cloning SauI-cleaved T. pallidum DNA into the cosmid pHC79. Sixteen of 800 clones investigated produced one or more antigens that reacted with antibodies from syphilitic patients. According to the separation pattern of the antigens produced on sodium dodecyl sulfate-polyacrylamide gels, six different phenotypes were distinguished among these 16 clones. These antigens reacted also with anti-T. pallidum rabbit serum. No antibodies against the cloned antigens were found in normal rabbit serum and in nonsyphilitic human serum. The antigens produced by the E. coli K-12 recombinant DNA clones comigrated in sodium dodecyl sulfate-polyacrylamide gels with antigens extracted from T. pallidum bacteria, suggesting that the treponemal DNA is well expressed in E. coli K-12. Several of the cosmid recombinant plasmids have been subcloned, resulting in smaller T. pallidum recombinant plasmids which are more stably maintained in the cell and produce more treponemal antigen. Monoclonal antibodies were raised against T. pallidum, and one hybridoma produced antibodies that reacted not only with an antigen from T. pallidum but also with the antigen produced by one of the E. coli clones.
|Keywords||*Cloning, Molecular, *Genes, Bacterial, 0 (Antigens, Bacterial), 0 (Genetic Vectors), 0 (Plasmids), Antigens, Bacterial/*genetics/immunology, Bacteriophage lambda, DNA Restriction Enzymes, EC 3.1.21 (DNA Restriction Enzymes), Escherichia coli/*genetics, Genetic Vectors, Plasmids, Treponema pallidum/*genetics|
van Embden, J.D.A., van de Donk, H.J.M., van Eijk, R.V.W., v.d. Heide, H.G., de Jong, J.A., van Olderen, M.F., … Schouls, L.M.. (1983). Molecular cloning and expression of Treponema pallidum DNA in Escherichia coli K12.. Infection and Immunity, 42(1), 187–196. Retrieved from http://hdl.handle.net/1765/3273