Team:TU-Delft/Notebook/2013/07/15/

The following Bio bricks from the distribution kit were transformed:

1. pBad araC agr BD - from IGEM agar stab received on 8/7/2013
2. RBS agrC RBS agrA TT pP2 - from IGEM agar stab received on 8/7/2013
3. RBS GFP TT
4. pTet RBS
5. cI
6. TT
7. RBS TetR TT
8. RBS RFP TT
9. pcI

The minipreps were performed to extract the plasmids.
Then the RBS agrC RBS agrA TT pP2 was linearized with SpeI and PstI and the RBS GFP TT was excised from its plasmid backbone using XbaI and PstI. The former was 4240 bp and the later was 876 bp. The digested products were run in the gel and eluted to do ligation:
RBS agrC RBS agrA TT pP2 + RBS GFP TT RBS agrC RBS agrA TT pP2: RBS GFP TT

The purpose for this ligation was to make an reporter AIP receiving strain of E. coli.
Then as a part of the timer construct the pTet RBS ( 2153 bp) was linearized with SpeI and PstI and the cI (776bp) was digested with XbaI and PstI. The two parts where ligated to get the pTet RBS: cI construct.
Some plasmids were linearised to facilitate fast working in the future. Those are as follows (numbers for figure 2)

* The insert and the excised backbone were almost same in length.
*Solution : A PCR reaction was set with the prefix and suffix primers and then the PCR product will be digested with X and P.
Gel pictures: