Team:HZAU-China/Attributions

Attributions

Our project is designed by Zhujun Xia and all circuits are designed by Zhujun Xia, YinQing Zeng and Mo Qiqin.

Wet Lab

Gene Editing

sifA and sipD knockout is completed by Zhujun Xia, Zhuoqi Huang and Wenxin Hu also gave assistance.

Plasmid Construction

Intracellular Environment-Dependent Circuits:

The intracellular environment-dependent circuits are designed by Mo Qiqin and Zhujun Xia.

The intracellular environment-dependent circuits are constructed by Mo Qiqin. Ruonan Tian also gave assistance.

Chemical Control System:

The ATc induction chemical control system is constructed by He Yang and Zhuoqi Huang also gave assistance.

The TetR-eGFP fusion protein expression system for TetR expression measurement is constructed by He Yang and 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 and Zhuoqi Huang by using 3A Assembly.

Plasmid Submission:

Submitted plasmids (Pcs2-eGFP-GSDMD FL, Pcs2-eGFP-GSDMD-N275 and two 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

Bacterial phenotype verifications are designed and completed by He Yang, including the OD measurement and CFU measurement of chemical control system.

Cell Phenotype Verification

Cell phenotype verifications are designed and completed by Zhujun Xia, including the verification of chemical control system, and targeting specificity, all the related circuits and the demonstration of the final work system.

Biosafety Verification

Safety verification is designed and completed by Zhujun Xia, which demonstrated our project is safe enough to be used.

InterLab

InterLab is conducted by Xichen Rao, Zhujun Xia, Ruonan Tian and Heng Heng also gave assistance. Our data passed the check of the Measurement Committee successfully.

Dry Lab

Mathematical model and app about salmonella infection and GSDMD release 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 on adjusting the strength of RBS and ATc with fixed GSDMD-N concentration is completed by Yini Miao.

The modeling of the promoter sifA and promoter entC is completed by Ruonan Tian.

Software

The applet of "Bacterial Colony Counter" is designed and programmed by Shuguang Wang.

Human Practice

Collaboration and communication with other teams are completed by Yinqing Zeng and all the other team members.

Handicraft manufacture is designed and completed by Yinqing Zeng.

Questionnaire preparation, the advertisement of synthetic biology and preparation of popular science articles are completed by Heng Heng.

Two posters for Eurasian exchange meetings and CCIC are completed by Ruonan Tian.

Website

Our wiki framework is designed and programmed by Shuguang Wang and our artist are Yuying Xiang and Kening Chen.

Our blog is built by Shuguang Wang.

Others

The plate reader machine operation is conducted by Mo Qiqin.

Chief cleaner of the laboratory is Xichen Rao.

Acknowledgements

For each parts of experiments, our appreciations are as follow:

Cell lines

Thanks to Prof. Shan Li’s laboratory, Huazhong Agricultural University, for sharing the cell lines of HEK293T, HELA, MCF7, iBMDM, so that our experiment can process successfully.

Thanks to Dr. Feng Shao’s laboratory, for sharing the cell line of GSDMD-/- HELA cells, which 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.

Bacteria

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 for generously sharing the material (Strain: E.coli K-12 MG1655 with mCherry in the genome).

Gene Knock In and Knock Out

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, which provides us with 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.

Plasmid Backbone

Thanks to Prof. Shan Li’s laboratory, Huazhong Agricultural University, for sharing the plasmid backbone (Pcs2-eGFP) to us.