Helping TUST prepare for their first year of competition.

To begin with, we offered them some constructive suggestions, and communicated with their team adviser. In 24/3/2017, we were invited to TUST to carried out a recruiting propaganda for them, and helped them form a team.

Helping NKU with their plasmid construction

What we did：
Helping NKU-iGEM construct their plasmid
The cleavage sites XbaI and SacI were added by PCR before and after lysase (lys)
Forward Primer：gcTCTAGAATGAAATACCTGCTGCCGAC
Reverse Primer：cGAGCTCtcaatgcgtttccataatagcagc

After construction

After the PCR product was obtained, then came cleavage：
Cutsmart® buffer 5μl
XbaI 1μl
SacI 1μl
PCR product 43μl
After overnight digestion, it was linking with the linearized plasmid pEX18 for 1 hr. Then the linking product was transformed into E.Coli.
PxylA-xylR was cleaved by SacI on plasmid pAX01
The pEX18-lys, which has been added with lys, was linearized by SacI single restrict digestion
The two were connected after 1hr
Then the linking product was transformed into E.coli.

E.coli after transformation

E.coli after transformation

Helped us to test the fluorescence intensity of pRS416-CUP1p-RFP without copper induction in Saccharomyces cerevisiae BY4742 and BY4741.
Cultures were incubated overnight in SC-URA medium.
Take some yeast suspension to the new SC-URA medium, then adjusted the OD600 value to 0.1 for 24 hours.
Fluorescence was measured, and the results were presented below:
Excitation 472nm
Radiation 532nm.

the data they obtained in their lab was provided for iGEM-Tianjin as reference

the data they obtained in their lab was provided for iGEM-Tianjin as reference

Test the mini system for OUC

What we did：

To verify whether the system built by China Ocean University was still available in other species of Saccharomyces cerevisiae.In that case, we used our laboratory-specific Saccharomyces cerevisiae with synthetic chromosome 10 to test its value of fluorescence intensity.

Protocol：

Yeast with plasmid was incubated overnight in YPD + G418 medium
Transfer the yeast suspension to the new YPD + G418 and adjusted the OD to 0.1
After incubation for 20 hours, the fluorescence was measured
Excitation light 502nm
Emitting light 532nm
The OD600 values were measured after fluorescence measurements

Having compared our results with that provided by Ocean University, except for some slight deviation of measurements, we found that the experimental results in both labs were consistent, which indicated that the mini system had similar expression in different laboratories and yeast strains.

Testing results for OUC

Testing results for OUC

What we asked OUC to do

Easy - to - error PCR library development. They were supposed to amplify our existing error-prone PCR library by conducting error-prone PCR for the CUP1 promoter we used in our project.
Specific steps:
1.Error-prone PCR
2. digestion
3. Purification / Adsorption
4. Connect
5. E.Coli transformation

Easy-to-error PCR protocol (100μl)：
5X buffer（140mM MgCl2, 250mM KCl, 50mM Tris, and 0.1%(wt/vol) gelatin）20μl
Template (iGEM-Tianjin provided) 4μl
Primers (iGEM-Tianjin provided) 4μl*2
10X dNTP (2mM dGTP, 2mM dATP, 10mM dCTP, and 10 mM TTP) 10μl
Taq polymerase 2μl
5mM MnCl2 10μl
ddH2O 46μl

94℃ 3min
94℃ 30s
53℃ 30s
72℃ 30s
72℃ 1min
recycle for 35 times

The PCR products were obtained and digested with BamHI and XbaI, and ligated with vector pRS416.
After E.Coli transformation, we collected the target bacteria and plasmids

Sent a Collaboration Request and construct an alliance to build a worldwide database.

We came up with the idea that we could gather all the iGEM team whose project is about water pollution treatment together and build an alliance to unite all our social impact, knowledge and geographical advantages.

2017.08.12

2. In 12th of August, we had a voice conferencing with SJTU/SCUT/XMU/UCAS/JLU/FAFU, we discussed about how we want to use this alliance and came up with 2 conclusion: 1. Build a worldwide database containing contents of heavy metals in soil 2. Mutual improve our social impact

2017.08.23

In 23th of August, we sent a Collaboration Request to iGEM official website, the next day Ana Sifuentes replied our message and posted our request in the iGEM official website,

and we received the response of many teams like team EXETER and team CSMU NCHU TAIWAN, with their information, we construct a database which contains the worldwide data of contents of Cu2+/Cd2+ in soil or water. And we build it based on map of the world. We received team CSMU NCHU TAIWAN’s kindly help, they offered us information about major metal pollution incidents in Taiwan as well as the real time monitoring data of places that have the potential to outbreak serious pollution incidents oneday.

Greenland

diagram 1

England

diagram 2

Australia

diagram 3

America

diagram 4

China

diagram 5

Africa

diagram 6

Russia

diagram 7

Helping CQU construct their team and sign up for their first year

We offered team CQU China many constructing advice including finding team advisor and allocating funds. Next yeat will be their first year joining iGEM, and we will continue helping them until they are ready for their Gamboree. Hope their result in next year will be great!

Filmed a biosafety video together with other 12 teams

12 teams gather together to film a biosafety video, every team took different topics, but all based on Yale biosafety manual.