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Team:TUDelft/1 September 2009
From 2009.igem.org
Lab Notebook
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1 September 2009
Sriram
Today I continued with the final assembly of Riboregulator. Since Daniel is leaving I also continued his assembly of locks and keys. Today I sent samples to sequenceing (Medium RBS control, Plasmid 1 of negative cascade, Plasmid 2 of negative cascade and Weak RBS Control). I added both primers given by TIm Weenink.
Daniel
Miniprep of yesterday's cultures:
| Part | [DNA] ng/uL |
| 1C | 8.5 |
| 2C | 45.6 |
| 3C | 20.5 |
| 4C | 16.6 |
| 1A | 161.2 |
| 1B | 84.9 |
| 2B | 95.2 |
| 3B | 265.1 |
| CP | 96.4 |
| RLacI | 58.3 |
As noticed, 1C concentration is low, this is because I add neutralization buffer before lysis buffer. We will do it again tomorrow.Thanks to Esengul, we will have electrocompetent cells which produce LacI.
Today I leave the lab and I'll be helping in planning and experimental design.
Tim Weenink
Today I restricted p(LacI) as an upstream part and *T1 (homing endonuclease without promotor) as a downstream part. Additionally I ligated the restricted I-SceI PCR fragment. This fragment was PCR'd from the plasmid from leiden containing the conding sequence of ISCEI. It was purified and restricted using E and P. Using the standard protocol, it was ligated into a pSB1A3 vector.
Finally, Esengul streaked a plate of K12 strain, that allows IPTG induction (without being expressed constitutively). This strain wil be grown up and made electro competent.
