Difference between revisions of "Team:Jiangnan/Demonstrate"
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<li><a href="https://2018.igem.org/Team:Jiangnan/Entrepreneurship">Gold</a></li> | <li><a href="https://2018.igem.org/Team:Jiangnan/Entrepreneurship">Gold</a></li> | ||
<li class="divider"></li> | <li class="divider"></li> | ||
| − | <li><a href="https://2018.igem.org/Team:Jiangnan/Public_Engagement"> | + | <li><a href="https://2018.igem.org/Team:Jiangnan/Public_Engagement">Public Engagement</a></li> |
<li class="divider"></li> | <li class="divider"></li> | ||
<li><a href="https://2018.igem.org/Team:Jiangnan/Entrepreneurship">Entrepreneurship</a></li> | <li><a href="https://2018.igem.org/Team:Jiangnan/Entrepreneurship">Entrepreneurship</a></li> | ||
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<div style="width:100%;margin-top:0px;padding-top:100px;background-color: #f0ebea"> | <div style="width:100%;margin-top:0px;padding-top:100px;background-color: #f0ebea"> | ||
| + | <img src="https://static.igem.org/mediawiki/2018/6/64/T--Jiangnan--demonstration_top.png" width="100%" style="margin-top: -35em;"> | ||
<div id="hightitration"> | <div id="hightitration"> | ||
<div class="row" style="position: relative;margin-top: 5em; margin-bottom: 5em;"> | <div class="row" style="position: relative;margin-top: 5em; margin-bottom: 5em;"> | ||
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<div style="float: left;width: 50%"> | <div style="float: left;width: 50%"> | ||
<img src="https://static.igem.org/mediawiki/2018/e/e5/T--Jiangnan--demonstration_high_figure1A.png" width="100%" height="650"> | <img src="https://static.igem.org/mediawiki/2018/e/e5/T--Jiangnan--demonstration_high_figure1A.png" width="100%" height="650"> | ||
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<p><b>Figure 1A.</b> IRF7 gene expression with different SiRNA concentration.</p> | <p><b>Figure 1A.</b> IRF7 gene expression with different SiRNA concentration.</p> | ||
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<div class="col s10 offset-s1"> | <div class="col s10 offset-s1"> | ||
<h5>2.Plasma </h5> | <h5>2.Plasma </h5> | ||
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<div class="col s11 offset-s1" style="text-align: center;"> | <div class="col s11 offset-s1" style="text-align: center;"> | ||
<img src="https://static.igem.org/mediawiki/2018/3/3d/T--Jiangnan--demonstration_high_figure2.png" width="60%"> | <img src="https://static.igem.org/mediawiki/2018/3/3d/T--Jiangnan--demonstration_high_figure2.png" width="60%"> | ||
| − | <p class="JFigure"><b>Figure 2</b> relative amount of viral DNA with different plasma treatment time.</p> | + | <p class="JFigure" style="font-size:0.8em;"><b>Figure 2</b> relative amount of viral DNA with different plasma treatment time.</p> |
<p>After incubating IRF7 silenced cells with the DMEM medium for 1 hour, IBRV replication in MDBK was further increased 2.5 folds. </p> | <p>After incubating IRF7 silenced cells with the DMEM medium for 1 hour, IBRV replication in MDBK was further increased 2.5 folds. </p> | ||
</div> | </div> | ||
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<div class="col s5" style="height: 800px;"> | <div class="col s5" style="height: 800px;"> | ||
| − | <img src="https://static.igem.org/mediawiki/2018/ | + | <img src="https://static.igem.org/mediawiki/2018/c/cc/T--Jiangnan--suspension_Jiangnan.png" width="100%" height="500"> |
| − | <p class="JFigure"><b>Figure 1</b> MA map. The X-axis was the mean value of all sample expression used for comparison after standardization, and the Y-axis was log2 transformed fold change. The significant p-value<0.05 was marked in red.</p> | + | <p class="JFigure" style="font-size:0.8em;"><b>Figure 1</b> MA map. The X-axis was the mean value of all sample expression used for comparison after standardization, and the Y-axis was log2 transformed fold change. The significant p-value<0.05 was marked in red.</p> |
<p>Genes differentially expressed between adherent and suspended cells were mapped to KEGG pathways.</p> | <p>Genes differentially expressed between adherent and suspended cells were mapped to KEGG pathways.</p> | ||
</div> | </div> | ||
<div class="col s5" style="height: 800px"> | <div class="col s5" style="height: 800px"> | ||
<img src="https://static.igem.org/mediawiki/2018/4/46/T--Jiangnan--demonstration_susp_figure2.png" width="100%" height="500"> | <img src="https://static.igem.org/mediawiki/2018/4/46/T--Jiangnan--demonstration_susp_figure2.png" width="100%" height="500"> | ||
| − | <p class="JFigure"><b>Figure 2</b> KEGG pathway enrichment map.</p> | + | <p class="JFigure" style="font-size:0.8em;"><b>Figure 2</b> KEGG pathway enrichment map.</p> |
<p>Red blocks indicate up-regulated genes, green blocks mean down-regulated genes, and the yellow one means both up-regulated and down-regulated gene.</p> | <p>Red blocks indicate up-regulated genes, green blocks mean down-regulated genes, and the yellow one means both up-regulated and down-regulated gene.</p> | ||
</div> | </div> | ||
<div class="col s5"> | <div class="col s5"> | ||
<img src="https://static.igem.org/mediawiki/2018/0/0c/T--Jiangnan--demonstration_susp_figure3.png" width="100%" height="500"> | <img src="https://static.igem.org/mediawiki/2018/0/0c/T--Jiangnan--demonstration_susp_figure3.png" width="100%" height="500"> | ||
| − | <p class="JFigure"><b>Figure 3</b> Venn diagram showing the number of genes in each analytical cohort.</p> | + | <p class="JFigure" style="font-size:0.8em;"><b>Figure 3</b> Venn diagram showing the number of genes in each analytical cohort.</p> |
</div> | </div> | ||
<div class="col s5"> | <div class="col s5"> | ||
<img src="https://static.igem.org/mediawiki/2018/3/34/T--Jiangnan--demonstration_susp_figure4.png" width="100%" height="500"> | <img src="https://static.igem.org/mediawiki/2018/3/34/T--Jiangnan--demonstration_susp_figure4.png" width="100%" height="500"> | ||
| − | <p class="JFigure"><b>Figure 4</b> Network showing interactions among candidate genes responsible for cell suspension feature. The 18 candidate genes were shown in the inner circle and other genes relevant with these candidates were retrieved by GeneMANIA and shown in the outer circle.</p> | + | <p class="JFigure" style="font-size:0.8em;"><b>Figure 4</b> Network showing interactions among candidate genes responsible for cell suspension feature. The 18 candidate genes were shown in the inner circle and other genes relevant with these candidates were retrieved by GeneMANIA and shown in the outer circle.</p> |
</div> | </div> | ||
<div class="col s10"> | <div class="col s10"> | ||
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<div class="col s5 offset-s4 JFigure"> | <div class="col s5 offset-s4 JFigure"> | ||
<img src="https://static.igem.org/mediawiki/2018/d/d9/T--Jiangnan--demonstration_susp_figure5.png" width="100%"> | <img src="https://static.igem.org/mediawiki/2018/d/d9/T--Jiangnan--demonstration_susp_figure5.png" width="100%"> | ||
| − | <p><b>Figure 5.</b></p> | + | <p><b>Figure 5.</b>Gene expression of PABPC1 in BHK-21 adherent and suspended cells.</p> |
</div> | </div> | ||
<div class="col s5 offset-s4"> | <div class="col s5 offset-s4"> | ||
| − | <p>We extracted RNA from BHK-21 adherent cells and suspension cells, and examined PABPC1 expressions in these cells by qPCR | + | <p>We extracted RNA from BHK-21 adherent cells and suspension cells, and examined PABPC1 expressions in these cells by qPCR.PABPC1 level in BHK-21 suspended cells is 0.4 times of that in BHK-21 adherent cells.</p> |
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</div> | </div> | ||
<div class="col s1" style="position: absolute;top :0px;padding-left: 0px;"> | <div class="col s1" style="position: absolute;top :0px;padding-left: 0px;"> | ||
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<div class="col s8 offset-s3" style="text-align: right;"> | <div class="col s8 offset-s3" style="text-align: right;"> | ||
<h5>3. siRNA transfection</h5> | <h5>3. siRNA transfection</h5> | ||
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</div> | </div> | ||
</div> | </div> | ||
| − | <div class="row" style="position: relative;margin-top: 5em;margin-bottom: | + | <div class="row" style="position: relative;margin-top: 5em;margin-bottom: 0;"> |
<div class="col s8 push-s3" style="text-align: right;"> | <div class="col s8 push-s3" style="text-align: right;"> | ||
<h5>5. Nectin 4 and viral coat protein expression detection by Indirect Immunofluorescence</h5> | <h5>5. Nectin 4 and viral coat protein expression detection by Indirect Immunofluorescence</h5> | ||
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<div class="col s8 offset-s2 JFigure" > | <div class="col s8 offset-s2 JFigure" > | ||
<img src="https://static.igem.org/mediawiki/2018/d/dd/T--Jiangnan--demonstration_broad_figure8.png" width="90%"> | <img src="https://static.igem.org/mediawiki/2018/d/dd/T--Jiangnan--demonstration_broad_figure8.png" width="90%"> | ||
| − | <p><b>Figure 9.< Envelope protein of CDV detected in MDBK-N4 cells by fluorescent microscope (×40).</p> | + | <p><b>Figure 9.</b> Envelope protein of CDV detected in MDBK-N4 cells by fluorescent microscope (×40).</p> |
<p>A: MDBK cells; B: MDBK-N4 cells</p> | <p>A: MDBK cells; B: MDBK-N4 cells</p> | ||
</div> | </div> | ||
Latest revision as of 03:03, 18 October 2018
<
1.IRF7
Though our computational modeling and network construction using public datasets retrieved from GEO（GSE76967，GSE32186，GSE40644，GSE8807，GSE8923，GSE3568） , we identified IRF7 as our candidate responsible for the high titration feature of cells. We increased virus titer over 2 folds by suppressing IRF7 expression in MDBK cells. In addition, by exposing IRF7-silenced MDBK cells to cold atmospheric plasma, we further increased virus titer to 2.5 fold. Taken, together, through genetic modification and physical plasma treatment, we could increase virus titer up to 5 folds.
Figure 1A. IRF7 gene expression with different SiRNA concentration.
Figure 1B.relative amount of viral DNA with different SiRNA concentration.
C: control NC: negative control
SiRF7-10nmol: SiRNA final concentration 10nmol/ml
SiRF7-50nmol: SiRNA final concnetration 50nmol/ml
Figure 1A showed that IRF7 was effectively knocked down, and Figure 1B showed that Bovine rhinotracheitis virus (IBRV) replication in the experimental group was over 2 folds than that in the negative control group after suppressing IRF7 expression (when siRNA concentration was 50nmol/ml).
2.Plasma
Figure 2 relative amount of viral DNA with different plasma treatment time.
After incubating IRF7 silenced cells with the DMEM medium for 1 hour, IBRV replication in MDBK was further increased 2.5 folds.
