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Team:KULeuven/6 August 2008
From 2008.igem.org
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Lab Work
Wet Lab
- The electrocompetent cells were tested with pUC and we had a lot of colonies so these are working great. *Ligation products were transformed to these electrocompetent cells (E0022+B0015, J23109+J23032, I712074+J23032, GFP+B0015, GFP+pSB1A2, C0062+B0015, pUC as a control).
- We made digest of J23032, F1610, J23022, B0015.
- Miniprep colonies of the ligation.
- The three punched parts of the memory gave a few colonies. One of them did not give any colonies at all. The colonies we had were plated on new plates.
Dry Lab
Check possibility of hybrid promoter to connect memory and timer (LuxI). This is to replace the antisense LuxI. Promotor would look something like this:
OperatorHSL_LuxR --- -35 box --- OperatorCII P22 --- -10 box
New modeling of the antisense LuxI and its target LuxI mRNA shows that this leaky repression is not a problem for the system. Still looking at the composite promoter though.
Hybrid promoter has been made! [http://partsregistry.org/Part:BBa_K145150 Hooray for BBa_K145150!] It should be activated by HSL-LuxR and repressed by P22 C2 (which is produced when the memory is OFF, hereby repressing LuxI synthesis) Check tomorrow how to best obtain the sequence.
Literature
[http://jb.asm.org/cgi/content/full/190/13/4392?view=long&pmid=18083819 Lux regulatory region (pLux)]
[http://www.jbc.org/cgi/content/full/272/3/1646/f1 P22 C2 repressor operators]
PS: P22 C2 is homologous to lambda CI while P22 C1 is homologous to lambda CII
Modeling
Wiki
Dr. Coli is everywhere, even at the URL.
Remarks
Discovery of the day. Two people beat us to the punch: [http://www.diagnosticinformationsystem.com/bios.html 1] and [http://www.sscofcny.com/doctors/coli.htm 2]
Sindsdien hangt uw haar slap -Maarten
