Team:Shenzhen BGIC ATCG/notes

Timeline

Promoter Group

Week1: research about the previous work of iGEM, especially the work done by the European teams.

Week2~week4: project idea discussing and determination， group issue asssigned.

Week5~week6: research about the principle of promoter in E.coli and yeast.

Week7: Experiments process design, constructs the basic parts and test curcuits.

Week8: Design the primers needed to amplification the promotor

Week9: Workshop among several schools

Week10-week13: constructs the basic parts: BBa_K1051300, BBa_K1051301, BBa_K1051302,BBa_K1051303, BBa_K1051304, BBa_K1051305, BBa_K1051306

Week14-week18: Measurement designing

Week19-week22: Test measurement circuits and then data analysis

Week23-week25: results drawing and made the wiki

Degradation Group

Week1: previous team project data collection, especially the Latin America teams

Week2~week4: project idea decided，group issue divided into E.coli one and yeast one.

Week5~week6: research of principle of degradation peptide and design the experiment generally combining with the mechanism of cyclin protein degradation.

Week7: Experiments process design, constructs the basic parts and test curcuits.

Week8: Design the primers needed to amplification the degradation tags.

Week9: Workshop among several schools

Week10-week13: constructs the basic parts: K051200/K1051201/K1051202/K1051203/K1051204/K1051205/K1051206/K1051207

Week14-week18: Measurement circuits construction

Week19-week22: Test measurement circuits and then data analysis

Week23-week25: results drawing and made the wiki

Targeting Group

Week1: research about the previous work of iGEM, especially the work done by the Asia teams.

Week2~week4: project idea discussing and determination， group issue assigned.

Week5~week6: research about the principle of promoter in E.coli and yeast.

Week7: Experiments process design, constructs the basic parts and test circuits.

Week8: Design the primers

Week9: Workshop among several schools

Week10-week13: constructs the basic parts: BBa_K1051100 to BBa_K1051118

Week14-week18: Measurement designing

Week19-week22: Test measurement circuits and then data analysis

Week23-week25: results drawing and made the wiki

Synchronization Group

Week1: research about the previous work of iGEM, especially the work done by the European teams.

Week2~week4: project idea discussing and determination， group issue assigned.

Week5~week6: research about the principle of promoter in E.coli and yeast.

Week7: Experiments process design, constructs the basic parts and test circuits.

Week8: Design the primers needed to amplification the degradation tags.

Week9: Workshop among several schools

Week10-week13: constructs the basic parts: BBa_K1051500, etc.

Week14-week18: Measurement designing

Week19-week22: Test measurement circuits and then data analysis

Week23-week25: results processing

Modeling

Week 1-8 research about the previous best model

Week 9 Decided using the budding yeast cell cycle model as basic part

Week 10-13 Learning the cell designer，Matlab Simbiology as the modeling software and trying to make some test

model.

Week 14-16 Understanding the cell cycle model in cell designer and using cell designer drawing reaction network,

setting the parameters.

Week 17 Learning how to use the Matlab Simbiology part and input results within cell designer

Week 18-20 Decided making three modeling: alternative splicing, Sic1 regulation and degradation tags. Do related research.

Week 21-23 Made the cell cycle circuits, find parameters

Week 24-25 combine with experiments data and made wiki

Attributions

Work Design

Gone Jianhui as a team leader and K2 as our instructor draft our project "Cell Magic".

Experiments Conduct

Li Xiang Li,Xu Yanhui,Wu Fanzi, Yu yang from SCU: responsible for targeting peptide,XFP,terminators design and experiments.

Chen Shihong,Gu Chenguang, Lu Yanping, Liang Jiale,from South China University of Technology, are responsible for the alternative splicing Src1 and Mer2 intron design and experiments.

Zhu Shuang, Lin Kequan from Wuhan University and Wei Wei, Zheng Bingwei, Yi Lan in HUST work together for the promotors.

Guan Rui from SEU and He funan, Wang Rui, Lin Li,Zhang Yaolei from UESTC made their efforts to the degradation parts.

Zhou Wanling,Zhang Aiping, Li Dongdong, the undergraduates in AHMU, joined the part one

Chen Yichun of SCNU, Zhong Na of JNU work for the microfulidic part.

The SCNU student: Chen Chengxuan, Lin Qiongfen, Xie Qiaolin, worked for the cell cycle regulator Sic1

Modeling

Liu Shuang Liu from SEU, Zhou Yang from SCUT, Jinchun Zhang from SCU, Qiu Bitao from BGI

Wiki Construction

Zhang Jinchun and Zhou Yang

Protocols

Protocol of microfluidic

1、 design the pattern of the biochip.

2、 print the pattern on the mask.

3、 prepare the basic materials like clean silicon wafer, photoresist and so on.

4、 spread the photoresist on the silicon wafer and make sure the thickness is 10 um.

5、 with the help of the ultraviolet ray, the pattern of the mask can be transferred to the metamorphic photoresist.

6、 (photographic fixing)Bake the silicon wafer to solidify the photoresist for about 2 hours.

7、 (develop the pattern)wash off the photoresist which wasn’t exposure to the ultraviolet ray by using the special chemical reagent

8、 Place the silicon wafer in the culture dish, then prepare the PDMS and pour it into the culture dish.

9、 When the PDMS freeze, downcut it and fit it on a slide by surface plasma.

10、 Connect the capillary tube and the chip.

Protocol of chip-based XFP degradation rate detection in E.coli.

First of all, E.coli will be measured after shaking to about OD2.0（600）in the chip which was washed by plasma water, vacuum pumping. The bacteria liquid was pushed into the chip, letting the cells enter the small triangle. Then use the constant flow pump culture medium into chip (the laboratory constant temperature, can not guarantee the training environment, 37°E. coli slower growth). The medium speed is about 200ul/h. Finally we test the data after yeast fulled in the triangles.Using the IPTG medium, the new RFP expression was stopped and we can regard the lights as degradation tags' efficiency.

Protocol of Enzyme - labelled meter detecting the fluorescent protein intensity

First of all, take a certain amount of bacteria liquid, recovery to around OD0.6(600), ensuring the bacteria in the logarithmic growth phase. Diluted and then the bacteria transferred to 96 well plates, measured their fluorescence intensity. Red fluorescent protein using an excitation wavelength of 584nm, and its emission wavelength is 607nm.When measuring, we first detect the OD600 of each strain, removing the factor of bacteria number difference. Thus the fluorescence intensity cannot be altered by bacteria quantity. Then the measurement mode switching for measurement of fluorescence, fluorescence intensity.

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Acknowledgements

At the Beginning

The BGI-ATCG iGEM 2013 team is a huge and diversity group consists of undergraduate from ten universities. Our team members’ major is really different, such as biotechnology bioinformatics, physical and even mathematics. Our work is partly assigned into each group made up by the students from the same university, which you can find in the group part in this page. And you can also browse about who and how everyone contributed to the project in this page. We really appreciate all our advisors and instructors that have assisted us throughout this project, without whom the project could not been carried out. We would also like to thank all everyone else who has helped us to achieve our project through put up advice or providing DNA, seeds, or other materials. Their contributions have helped us enormously. For a full list of acknowledgments, please see the bottom of this page.

Financial Support

BGI College provides the totally funding including our team registration fee and competition travel fee.

SUSTC Biology Department and the BGI Unit of Synthetic Biology help us purchasing some materials.

General Support

Cho-Kiu Wong at SMC, BGI Tech Solutions help us much to send and receive the BioBricks

Unit of Synthetic Biology at BGI trains team members about the basic experiment technology and knowledge which was really necessary for us.

Material Support

Boeke Lab of John Hopkins Medical Institutions (Boeke Lab @ John Hopkins Medical) provide us pRS413, pRS414, pRS415, pRS416

Dr. Chi-Ming Wong (Dr. Chi-Ming Wong @ HKU) at Hongkong University provides us the plasmid YEpLac195 YEpLac181

Kai Tian and Yong Li at BGI, helping us with the CRIPSR system.

WHU-China has provided us the E. coli version of CRISPRi system.

At Last

We really appreciate 1) the BGIC_0101 team for their cooperation, 2) the SCSTC help us for borrowing device and booking materials, and 3) SYSU, SUSTC borrowing us some experiments equipment.

Lab Support

Unit of Synthetic Biology at BGI supports us with the lab for the most important cell and molecular biology experiments.

Dr. Ming Ni, team leader in BGI cancer group, let us use his lab for the microfluidic experiments.

Pr. Lingling Shui at south china normal university provides the microfluidic lab and materials for our experiments.

Cooperation College or University

Huazhong University of Science and Technology

Wuhan University

China University of Geosciences

South China University of Technology

South China Normal University

Jinan University

Sichuan University

University of Electronic Science and Technology of China

Southeast University

Qingdao University

University of Chinese Academy of Sciences