The production of strains highly expressing human interleukin-2 cDNA.
The plasmid pTLIL-2, which only directed a low-level expression of human interleukin 2(IL-2) cDNA in E. coli, was reconstructed: a series of deletions were made in 3' non-coding region of human IL-2 cDNA, and 7 recombinant plasmids with different deletions were selected, on the other hand, a Tac promoter sequence from pDR540 was inserted to upstream of IL-2 cDNA in pTLIL-2 so that pTLIL-2DT, which contains double Tac promoters, was constructed. And then, E. coli JM103 was transformed with the above 8 recombinant plasmids respectively. The expression efficiency of IL-2 cDNA in E. coli transformed with different plasmids was evaluated by means of SDS-PAGE and measuring 3H-TdR incorporation of IL-2-dependent activated T lymphocytes in the presence of bacterial extracts. Three engineering strains with high efficiency of IL-2 expression were selected, and all of these strains could produce recombinant IL-2(rIL-2) 4 times more than E. coil JM103/pTLIL-2.
Zhang
D J
DJ
Department of Medical Molecular Biology, Tongji Medical University, Wuhan.
Feng
B
B
Huangfu
Y M
YM
D004251:DNA Transposable Elements; D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D007376:Interleukin-2 / Q000096:biosynthesis* / Q000235:genetics; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis*; D013045:Species Specificity
D004251:DNA Transposable Elements; D018076:DNA, Complementary; D007376:Interleukin-2; D011994:Recombinant Proteins