Avaliação do sistema de estabilização plasmidial toxina-antitoxina para a produção de proteínas recombinantes em <i>Escherichia coli</i>.

Abstract

KATAYAMA, K.Y. Evaluation of plasmidial stabilization system toxin-antitoxin for recombinant proteins production in Escherichia coli. 2016. 66 p. Master thesis

Development and in silico evaluation of an expression platform based on E.coli for the production of a recombinant beta-glucosidase.

“…Values of total soluble proteins (obtained by the Bradford method), OD600 and dry cell weight for the same samples are gathered in Table 1212 as well, revealing that the SE1/BGL system produced a higher activity than the BL21/BGL system, in spite of similar levels of dry cell weight and total soluble proteins. These results concur with those of (189,264,266), who also observed higher expression levels for the SE1 toxin-antitoxin system than for the BL21 + antibiotic system. CodonPlus strain with a pET21a plasmid carrying the same BGL gene.…”

“…Nonetheless, plasmid stability is still very high for E. coli BL21 in the beginning of the assay -of 97% for 12 h, 24 h and 36 h -and decreases moderately to 92% at t = 60 h. These results corroborate other works in the literature. (189,264,266), which found that the SE1+pStaby system preserves plasmid stability at approximately 100%, as opposed to the BL21(DE3) + antibiotic system, which suffers from plasmid loss over time. Moreover, they validate the assumption that the SE1 strain would be ideal for the proposed inoculum production scheme.…”

“…As such, the sole differences between the strains are the additional genes ccdB and Cm R present in the SE1 strain; the ccdB gene gives rise to the ccdB toxin, while the Cm R gene provides resistance to chloramphenicol (an antibiotic not used in this work). Although no direct comparison of the two strains grown on defined medium was found in the literature other than the aforementioned ref (264),. the few works that do compare SE1 and BL21(189,266) have not found significant differences between the strains in terms of viable cell count or dry cell weight.…”