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Team:KU Leuven/Journal/FFL/wetlab
From 2013.igem.org
Secret garden
Congratulations! You've found our secret garden! Follow the instructions below and win a great prize at the World jamboree!
- A video shows that two of our team members are having great fun at our favourite company. Do you know the name of the second member that appears in the video?
- For one of our models we had to do very extensive computations. To prevent our own computers from overheating and to keep the temperature in our iGEM room at a normal level, we used a supercomputer. Which centre maintains this supercomputer? (Dutch abbreviation)
- We organised a symposium with a debate, some seminars and 2 iGEM project presentations. An iGEM team came all the way from the Netherlands to present their project. What is the name of their city?
Now put all of these in this URL:https://2013.igem.org/Team:KU_Leuven/(firstname)(abbreviation)(city), (loose the brackets and put everything in lowercase) and follow the very last instruction to get your special jamboree prize!
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Week4: Getting started
This week we started working on the proof of concept of the feed forward loop in the lab. First of all we selected the necessary bricks and took the plasmids out of the iGEM distribution kit plates. We transformed them into chemically competent inoue cells (which were made before and stored at -80°C) according to the ‘transformation kit protocol’.
Following bricks were transformed: C0076, K823016, C0077, R0080, P0340, R0078, K082003, C0080 and R0011.
All of them were plated out in the presence of chloramphenicol, except for R0011 which is resistant to ampicilline. We made 2 plates for every brick, one with 100 µl and one with 1000 µl LB. The bacteria were grown overnight at 37°C.
The next day we checked our plates and all of them had colonies except for the plates of R0011. We picked one colony of every plate and put the bacteria into tubes with 4 ml of LB medium and 4 µL chloramfenicol to incubate overnight in a shaking incubator at 37°C. Then we decided to transform the failed brick (R0011) and an extra brick (C0060) by electroporation because the efficiency of this method should be higher. We plated them out and let them grow overnight at 37°C.
The plates with brick C0060 had nicely grown colonies and we put them at 4°C for usage next week. The plates with brick R0011 didn't show colonies. Afterwards we noticed that we used the wrong antibiotic resistance. We took the tubes with the successful bricks out of the shaking incubator and we used 400 µl of the LB with bacteria together with 400 µl of glycerol to make a stock in the -80°C.
The remaining 3.6 ml was used for a plasmid extraction using the home-made protocol. After the extraction we used the NanoDrop to see if we had a good amount of DNA and no interference of proteins or salts. The values are shown below:ng/µl 260/280 260/230 C0076 A 94.6 1.34 0.55 B 10.9 1.37 0.41 K823016 A 12.5 1.29 0.43 B 8.7 1.34 0.38 C0077 A 4.1 1.91 0.48 B 2.9 1.70 0.46 R0080 A 18.1 2.02 0.71 B 4.9 1.23 0.32 P0340 A 73.3 1.83 0.99 B 155.8 1.92 1.52 R0078 111.6 1.73 1.07 K082003 A 127.3 1.89 1.55 B 197.1 2.11 1.29 C0080 A 200.6 1.81 1.24 B 7.2 1.92 1.04 -
We missed Tomas!
Tomas, the man behind our ecological model has been on holiday. Now he's back and ready to finetune his diffusion model.
