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<div class="h3">Plasmid Construction</div> | <div class="h3">Plasmid Construction</div> | ||
<p>Intracellular Environment-Dependent Circuits:</p> | <p>Intracellular Environment-Dependent Circuits:</p> | ||
− | <p>The intracellular environment-dependent circuits is designed by Mo | + | <p>The intracellular environment-dependent circuits is designed by Mo Qiqin and Zhujun Xia.</p> |
<p>The intracellular environment-dependent circuits is constructed by Mo Qiqin. Ruonan Tian also gave | <p>The intracellular environment-dependent circuits is constructed by Mo Qiqin. Ruonan Tian also gave | ||
assistance.</p> | assistance.</p> | ||
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<br> | <br> | ||
<p>Targeting Specificity and Surface-Displaying:</p> | <p>Targeting Specificity and Surface-Displaying:</p> | ||
− | <p>RGD-inserted OmpA circuit clone and construction of Salmonella SL1344_RS05255 (OmpA coding | + | <p>RGD-inserted OmpA circuit clone and construction of <i>Salmonella</i> SL1344_RS05255 (OmpA coding |
sequence) are completed by Yinqing Zeng and Lingyu Zhong.</p> | sequence) are completed by Yinqing Zeng and Lingyu Zhong.</p> | ||
<p>The circuit of surface-display system is constructed by Lingyu Zhong. | <p>The circuit of surface-display system is constructed by Lingyu Zhong. | ||
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<p>Submitted plasmids (Pcs2-eGFP-GSDMD FL, Pcs2-eGFP-GSDMD-N275 and three GSDMD full length mutations) | <p>Submitted plasmids (Pcs2-eGFP-GSDMD FL, Pcs2-eGFP-GSDMD-N275 and three GSDMD full length mutations) | ||
are constructed by Zhiqing Guo and Wenxin Hu.</p> | are constructed by Zhiqing Guo and Wenxin Hu.</p> | ||
− | <p>The description of submitted plasmids by ATP assay and cell phenotype via transfection is | + | <p>The description of submitted plasmids by ATP assay and cell phenotype via transfection is designed |
and completed by Zhujun Xia. | and completed by Zhujun Xia. | ||
</p> | </p> | ||
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and CFU measurement of chemical control system.</p> | and CFU measurement of chemical control system.</p> | ||
<div class="h3">Cell Phenotype Verification</div> | <div class="h3">Cell Phenotype Verification</div> | ||
− | <p>Cell phenotype verifications are designed and completed by Zhujun Xia, including the | + | <p>Cell phenotype verifications are designed and completed by Zhujun Xia, including the verification |
of chemical control system, and targeting specificity, all these circuits and systems work so well.</p> | of chemical control system, and targeting specificity, all these circuits and systems work so well.</p> | ||
<div class="h3">Bio-Safety Verification</div> | <div class="h3">Bio-Safety Verification</div> | ||
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our data passed the check of the Measurement Committee successfully.</p> | our data passed the check of the Measurement Committee successfully.</p> | ||
<div class="h2">Dry Lab</div> | <div class="h2">Dry Lab</div> | ||
− | <p>Mathematical model and app of | + | <p>Mathematical model and app of <i>Salmonella</i> infection and the releasing of GSDMD are built by Songtao |
Chen. | Chen. | ||
ATc model which predicted the relationship between concentration of ATc and concentration of GSDMD | ATc model which predicted the relationship between concentration of ATc and concentration of GSDMD | ||
is built by Xichen Rao. | is built by Xichen Rao. | ||
− | The modeling of adjusting the strength of RBS and ATc with fixed GSDMD- | + | The modeling of adjusting the strength of RBS and ATc with fixed GSDMD-N275 concentration is completed |
by Yini Miao. The future work is confirming the threshold of GSDMD-N275. | by Yini Miao. The future work is confirming the threshold of GSDMD-N275. | ||
The modeling of the promoter sifA and promoter entC is completed by Ruonan Tian. | The modeling of the promoter sifA and promoter entC is completed by Ruonan Tian. | ||
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of HEK293T, HELA, MCF7, iBMDM, our experiment can process successfully. | of HEK293T, HELA, MCF7, iBMDM, our experiment can process successfully. | ||
</p> | </p> | ||
− | <p> Thanks to Dr. Feng | + | <p> Thanks to Dr. Feng Shao's laboratory, for sharing the cell line of GSDMD<sup>-/-</sup> HELA cells, it provides |
us great support. | us great support. | ||
</p> | </p> | ||
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<div class="h2">Bacteria</div> | <div class="h2">Bacteria</div> | ||
<p>Thanks to Prof. Shan Li’s laboratory, Huazhong Agricultural University, for sharing the strain of | <p>Thanks to Prof. Shan Li’s laboratory, Huazhong Agricultural University, for sharing the strain of | ||
− | Salmonella enterica var. Typhimurium SL1344. | + | <i>Salmonella</i> <i>enterica</i> var. Typhimurium SL1344. |
</p> | </p> | ||
− | <p> Thanks to Dr. Chenli Liu from SIAT CSynBER’s generous share (Strain: E.coli K-12 MG1655 with | + | <p> Thanks to Dr. Chenli Liu from SIAT CSynBER’s generous share (Strain: <i>E.coli</i> K-12 MG1655 with |
mCherry in the genome), our experiment can process successfully. | mCherry in the genome), our experiment can process successfully. | ||
</p> | </p> |
Revision as of 12:38, 15 October 2018
Our project is designed by Zhujun Xia and all circuits is designed by Zhujun Xia, YinQing Zeng and Mo Qiqin.
SifA knock out is completed by Zhunjun Xia. Zhuoqi Huang and Wenxin Hu also gave assistance.
Intracellular Environment-Dependent Circuits:
The intracellular environment-dependent circuits is designed by Mo Qiqin and Zhujun Xia.
The intracellular environment-dependent circuits is constructed by Mo Qiqin. Ruonan Tian also gave assistance.
Chemical Control System:
The ATc induced chemical control system is constructed by He Yang, Zhuoqi Huang also gave assistance.
The TetR-eGFP fusion protein expression system for TetR expression measuring is constructed by He Yang, Zhuoqi Huang also gave assistance.
Targeting Specificity and Surface-Displaying:
RGD-inserted OmpA circuit clone and construction of Salmonella SL1344_RS05255 (OmpA coding sequence) are completed by Yinqing Zeng and Lingyu Zhong.
The circuit of surface-display system is constructed by Lingyu Zhong. Optimation of OmpA in E. Coli is completed by Zhujun Xia.
Plasmid Standardization:
Plasmid standardization is completed by Zhiqing Guo, Zhuoqi Huang by using 3A Assembly.
Plasmid Submission:
Submitted plasmids (Pcs2-eGFP-GSDMD FL, Pcs2-eGFP-GSDMD-N275 and three GSDMD full length mutations) are constructed by Zhiqing Guo and Wenxin Hu.
The description of submitted plasmids by ATP assay and cell phenotype via transfection is designed and completed by Zhujun Xia.
Bacterial phenotype verification are designed and completed by He Yang, including the OD measurement and CFU measurement of chemical control system.
Cell phenotype verifications are designed and completed by Zhujun Xia, including the verification of chemical control system, and targeting specificity, all these circuits and systems work so well.
Safety verification is designed and completed by Zhujun Xia, which demonstrated our project is safe enough to be used.
InterLab is conducted by Xichen Rao, Zhunjun Xia, Ruonan Tian and Heng Heng also gave assistance, our data passed the check of the Measurement Committee successfully.
Mathematical model and app of Salmonella infection and the releasing of GSDMD are built by Songtao Chen. ATc model which predicted the relationship between concentration of ATc and concentration of GSDMD is built by Xichen Rao. The modeling of adjusting the strength of RBS and ATc with fixed GSDMD-N275 concentration is completed by Yini Miao. The future work is confirming the threshold of GSDMD-N275. The modeling of the promoter sifA and promoter entC is completed by Ruonan Tian.
The applet of “Bacterial Colony Counter” is designed and programmed by Shuguang Wang.
Collaboration and communication with other teams is completed by Yinqing Zeng. Handicraft manufacture is designed and completed by Yinqing Zeng. Questionnaire preparation, the advertisement of synthetic biology and popular science articles written is completed by Heng Heng. Two posters for Eurasian exchange meetings and CCIC is completed by Ruonan Tian.
Our intergrative wiki is designed and programmed by Shuguang Wang and the artist is designed by Yuying Xiang.
Our blog is built by Shuguang Wang.
The plate reader machine operation is conducted by Mo Qiqin. Chief cleaner of the laboratory is Xichen Rao.
For each parts of experiments, our appreciations are as follow:
Thanks to Prof. Shan Li’s laboratory, Huazhong Agricultural University, for sharing the cell lines of HEK293T, HELA, MCF7, iBMDM, our experiment can process successfully.
Thanks to Dr. Feng Shao's laboratory, for sharing the cell line of GSDMD-/- HELA cells, it provides us great support.
Thanks to Dr. Yi Zeng and Dr. Ms Hong Fan, Shanghai Institute of Biochemistry and Cell Biology, CAS, for sharing the cell line of MDA-MB-231.
Thanks to Prof. Shan Li’s laboratory, Huazhong Agricultural University, for sharing the strain of Salmonella enterica var. Typhimurium SL1344.
Thanks to Dr. Chenli Liu from SIAT CSynBER’s generous share (Strain: E.coli K-12 MG1655 with mCherry in the genome), our experiment can process successfully.
Thanks to Prof. Shan Li’s laboratory, Huazhong Agricultural University, for providing the protocol and material of gene editing based on two-step allelic exchange, it provides us great help.
Thanks to Pan Chu, team leader of iGEM-2016, Huazhong Agricultural University, for teaching us multigene editing based on CRISPR and λ-RED.
Thanks to Prof. Shan Li’s laboratory, Huazhong Agricultural University, for sharing the plasmid backbone (Pcs2-eGFP) to us.
Thanks to Genescript and IDT for DNA sequence synthesis service
Thanks to Snapgene and MathWorks for free software supporting
Thanks to College of Life Science and technology of Huazhong Agricultural University for providing the site and fund for us.
Thanks to Prof. Binguang Ma for instructing our modeling.
Thanks to Prof. Jin He as our Secondary PI and Prof. Shan Li for instructing our experiment.
Thanks to Prof. Jin He, Prof. Shan Li, Prof. Gang Cao, Prof. Donghai Peng and Prof. Ming Sun for giving us advices at the proposal presentation.
Thanks to Kening Chen, Wenqi Huang and Haimeng Li from iGEM-2017, Huazhong Agricultural University, for giving us advices.
We really appreciate your support!