Designed by: Shuangshuang Pu Group: iGEM18_LZU-CHINA (2018-10-05)
HRE(Hypoxia response elements)-miniCMV
- In solid tuumors, rapid proliferation of tumor cells leads to increased oxygen consumption of tumor cells, which in turn leads to hypoxia microenviroment. Hypoxia is one of the basic characteristics of tumor microenvironment, playing an important role in the proliferation, invasion and metastasis of tumor cells. In our experiments, we transfect 293T cell with HRE-miniCMV and when it senses the condition of hypoxia in tumor microenvironment, HIF-1(hypoxia inducible factor 1) is produced, and it can bind to HRE(hypoxia response elements), then activating the minimized CMV promoter. Through this mechanism, we can switch the interest gene specifically in tumor microenvironment. In order to enhance its induction effect,we designed five HRE repeats components.After pHRE, miniCMV was added to transcribe the downstream genes.
- According to the experimental results(Figure 1 and 2), HRE promoter can induce higher copGFP fluorescence level under hypoxia conditions.But at high concentrations of CoCl(II), cells apoptosis as a result of metal toxicity, leading to a sharp decrease in fluorescence intensity.The overall fluorescence level of gastrical cancer cell MGC803 was lower than that of 293T(Figure 2), which may be due to high degree differentiation of MGC803,leading to an unstable expression level.
- 10COMPATIBLE WITH RFC
- 12COMPATIBLE WITH RFC
- 21Illegal XhoI site found at 5
- 23COMPATIBLE WITH RFC
- 25COMPATIBLE WITH RFC
- 1000COMPATIBLE WITH RFC
- The functions of five repeating HRE components were verified through CopGFP.The mechanism of hypoxia induced regulation is shown in the above figure:
Sequence and Features