[Construction of Escherichia coli-Mycobacteria shuttle plasmid and the stable expression of human interleukin-2 in BCG and Escherichia coli].

OBJECTIVE

To construct and identify Escherchia coli (E. coli)-Mycobacteria shuttle plasmid and to detect stable expression of foreign gene in E. coli and BCG.

METHOD

With a genetic engineering technique to construct the E. coli-Mycobacteria shuttle plasmid, the human interleukin-2 (IL-2) gene was electrophoreted into BCG with recombinant plasmid PZSIII-I and positive clones selected using polymerase chain reaction (PCR) technique. The expression of foreign gene of human IL-2 in BCG was identified by ELISA and SDS-PAGE.

RESULT

Human IL-2 cytokine was steadily expressed in recombinant BCG and E. coli and could secrete outside the cell.

CONCLUSION

M. bovis BCG recombinant constructed can produce and secrete the human IL-2. A secretion of the active cytokine was accomplished through the combined use of the BCG HSP65 promoter and a secretion signal from the BCG Ag-85B. The BCG HSP65 promoter is active in both BCG as well as E. coli which can not secrete foreign protein.