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.cebian { | .cebian { | ||
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font-family: 'Product Sans', sans-serif; | font-family: 'Product Sans', sans-serif; | ||
font-size: 40px; | font-size: 40px; | ||
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font-weight: bold; | font-weight: bold; | ||
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font-family: 'Product Sans', sans-serif; | font-family: 'Product Sans', sans-serif; | ||
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line-height: 30px; | line-height: 30px; | ||
font-weight: bold; | font-weight: bold; | ||
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font-family: 'Product Sans', sans-serif; | font-family: 'Product Sans', sans-serif; | ||
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+ | /* text-align: left !important; */ | ||
color: #ffffff !important; | color: #ffffff !important; | ||
font-size: 16px; | font-size: 16px; | ||
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background-color: #323643; | background-color: #323643; | ||
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font-family: Arial; | font-family: Arial; | ||
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color: #676767; | color: #676767; | ||
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+ | .cebian { | ||
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+ | .zhengwen { | ||
+ | margin: 20px 20px 0 0; | ||
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.daohangyi { | .daohangyi { | ||
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+ | .daohangyi img { | ||
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− | <p | + | <p>Our project goal is to induce pyroptosis in tumor cells through |
− | + | translocating a pyroptosis-associated protein GSDMD by <i>Salmonella</i>. Expression of the N-terminal of | |
− | + | GSDMD fused with eGFP (eGFP-GSDMD-N275) is under the control of the promoter P<sub>tet</sub> in Δ<i>sifA</i> | |
− | + | Salmonella. Hela GSDMD KO (knockout) cell line were infected with Δ<i>sifA</i> <i>Salmonella</i>. Inducer anhydrotetracycline (ATc) (16μg/mL) was added 3 h after infection. Microscopy shows that eGFP-GSDMD-N275 located in cytoplasm after 5 min of induction and pyroptosis was triggered after 30 min of induction (<b>Figure 1</b>). By counting the population of ruptured cells, a 1.96 fold-change was demonstrated between the induced group and the control group (<b>Figure 2</b>). So, the pyroptosis of host cell in the induced group was triggered by eGFP-GSDMD-N275 as expected. These results demonstrate that we successfully achieved the goal of our project!!! </p> | |
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− | GSDMD fused with eGFP (eGFP-GSDMD-N275) is under the control of P<sub>tet</sub> in Δ<i>sifA</i> | + | |
− | Salmonella. Hela GSDMD KO cell line were | + | |
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<img src="https://static.igem.org/mediawiki/2018/b/b3/T--HZAU-China--basicPart4.png" width=100% alt=""> | <img src="https://static.igem.org/mediawiki/2018/b/b3/T--HZAU-China--basicPart4.png" width=100% alt=""> | ||
</div> | </div> | ||
− | <p>Figure 1. Hela GSDMD KO | + | <p><b>Figure 1.</b> Hela GSDMD KO cells were infected by <i>Salmonella</i> Δ<i>sifA</i> SL1344 containing high copy number plasmids which express eGFP-GSDMD-N275 under the control of ATc. Photographs were captured 5 min, 30 min, and 90 min after induction, respectively. </p> |
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<div style="width: 40%; margin: 30px auto"> | <div style="width: 40%; margin: 30px auto"> | ||
<img src="https://static.igem.org/mediawiki/2018/1/16/T--HZAU-China--Demonstrate2.png" width=100% alt=""> | <img src="https://static.igem.org/mediawiki/2018/1/16/T--HZAU-China--Demonstrate2.png" width=100% alt=""> | ||
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− | <p>Figure 2. Ruptured cells in a field of view were counted (n=8). | + | <p><b>Figure 2.</b> Ruptured cells in a field of view were counted (n=8).</p><br> |
− | + | <p>In these experiments, the choice of MOI (multiplicity of infection) and infection time were directed by <a href="https://2018.igem.org/Team:HZAU-China/Model">Modeling</a>.</p> | |
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<!-- 侧边 --> | <!-- 侧边 --> |
Latest revision as of 02:41, 18 October 2018
Our project goal is to induce pyroptosis in tumor cells through translocating a pyroptosis-associated protein GSDMD by Salmonella. Expression of the N-terminal of GSDMD fused with eGFP (eGFP-GSDMD-N275) is under the control of the promoter Ptet in ΔsifA Salmonella. Hela GSDMD KO (knockout) cell line were infected with ΔsifA Salmonella. Inducer anhydrotetracycline (ATc) (16μg/mL) was added 3 h after infection. Microscopy shows that eGFP-GSDMD-N275 located in cytoplasm after 5 min of induction and pyroptosis was triggered after 30 min of induction (Figure 1). By counting the population of ruptured cells, a 1.96 fold-change was demonstrated between the induced group and the control group (Figure 2). So, the pyroptosis of host cell in the induced group was triggered by eGFP-GSDMD-N275 as expected. These results demonstrate that we successfully achieved the goal of our project!!!
![](https://static.igem.org/mediawiki/2018/b/b3/T--HZAU-China--basicPart4.png)
Figure 1. Hela GSDMD KO cells were infected by Salmonella ΔsifA SL1344 containing high copy number plasmids which express eGFP-GSDMD-N275 under the control of ATc. Photographs were captured 5 min, 30 min, and 90 min after induction, respectively.
![](https://static.igem.org/mediawiki/2018/1/16/T--HZAU-China--Demonstrate2.png)
Figure 2. Ruptured cells in a field of view were counted (n=8).
In these experiments, the choice of MOI (multiplicity of infection) and infection time were directed by Modeling.