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EPF-Lausanne/20 August 2009
From 2009.igem.org
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==Wet Lab== | ==Wet Lab== | ||
| + | Long day....... | ||
| + | Miniprep of the LovTap-Term clones we'd put in liquid culture. Then we tried to do the whole protocol to obtain our BioBrick LacI-RBS-LovTap-Term by 2 strategies: | ||
| + | |||
| + | - classic iGEM protocol: amplify LacI-RBS by PCR, purify, then digest it with E/S and digest the LovTap-Term vector with E/X , dephosphorylate the vector, purify again, then do the ligations of the various LacI-RBS with the LovTap-Terms in random combinations | ||
| + | |||
| + | - the "dirty" technique: digest both the LovTap-Term and LacI-RBS plasmids, dephosphorylate the LovTap-Term vector, purify, then do the ligation by just "mixing" the 2 solutions (so containing pieces of both vectors). The point is that since the LovTap-Term vector has a kanamycin resistance and the LacI-RBS is ampicilin, only the correct combination should manage to grow | ||
| + | |||
| + | We then did the transformations with the resulting ligation products. Plates were left to incubate overnight at 37°C. | ||
| + | |||
| + | Also, we obtained 2 clones from the previous day transformations with readout system 1. Did a colony PCR and a gel to check the insert, but the result of the gel showed some bands which we couldn't seem to identify. Did liquid cultures of the 2 clones so that we can do more tests tomorrow. | ||
==People in the lab== | ==People in the lab== | ||
| - | Basile, Gab, Christian | + | Basile, Gab, Christian, Heidi |
Latest revision as of 13:56, 21 August 2009
Contents |
Wet Lab
Long day.......
Miniprep of the LovTap-Term clones we'd put in liquid culture. Then we tried to do the whole protocol to obtain our BioBrick LacI-RBS-LovTap-Term by 2 strategies:
- classic iGEM protocol: amplify LacI-RBS by PCR, purify, then digest it with E/S and digest the LovTap-Term vector with E/X , dephosphorylate the vector, purify again, then do the ligations of the various LacI-RBS with the LovTap-Terms in random combinations
- the "dirty" technique: digest both the LovTap-Term and LacI-RBS plasmids, dephosphorylate the LovTap-Term vector, purify, then do the ligation by just "mixing" the 2 solutions (so containing pieces of both vectors). The point is that since the LovTap-Term vector has a kanamycin resistance and the LacI-RBS is ampicilin, only the correct combination should manage to grow
We then did the transformations with the resulting ligation products. Plates were left to incubate overnight at 37°C.
Also, we obtained 2 clones from the previous day transformations with readout system 1. Did a colony PCR and a gel to check the insert, but the result of the gel showed some bands which we couldn't seem to identify. Did liquid cultures of the 2 clones so that we can do more tests tomorrow.
People in the lab
Basile, Gab, Christian, Heidi
