Difference between revisions of "Part:BBa K2817002"

Latest revision as of 17:35, 17 October 2018

IL10-flag

Interleukin-10 is a natural immunosuppressive cytokine that is normally released by regulatory T cells and plays an important role in intestinal homeostasis. In our engineered bacteria, we hope to induce the release of interleukin-10 through the inflammatory signal nitric oxide to achieve drug release controllability.

Usage and Biology

We inserted this part into plasmid pCDFDuet-1. So the expression of this part can be induced by IPTG. We transformed the human IL10-Flag plasmid into BL21, incubated at 37 ℃ overnight and then diluted to OD600 = 0.2. After 2 h of growth at 37 ℃, IPTG was added to induce IL10 expression at 30 ℃ for 16 h. Then, the bacterial solution was lysed and the expressed IL10 was detected by Western Blot (Figure 1). From this result, IL10 was successfully expressed in our constructed plasmid. As a widely validated and used anti-inflammatory interleukin, IL10 has been identified with critical functions in immunosuppression in human [1].

Figure 1. Western blot analyses using a flag antibody on bacterial lysate to detect IL10.

Lane 1: Negative control (cell lysate without IPTG induction); Lane 2: 0.5mM IPTG.

[1] Steidler, L. Treatment of murine colitis by Lactococcus lactis secreting interleukin-10. Science 289, 1352–1355 (2000).

Sequence and Features