http://2013.igem.org/wiki/index.php?title=Team:Heidelberg/Templates/Indigoidine_week5&feed=atom&action=historyTeam:Heidelberg/Templates/Indigoidine week5 - Revision history2024-03-28T12:06:06ZRevision history for this page on the wikiMediaWiki 1.16.5http://2013.igem.org/wiki/index.php?title=Team:Heidelberg/Templates/Indigoidine_week5&diff=310515&oldid=prevFloSmi: Created page with " We got two plasmids from the Fussenegger group at the ETH Zurich containing a codon optimized version of bpsA (pMM64) and svp (pMM65), respectively, for expression in HEK 297-..."2013-10-05T00:28:07Z<p>Created page with " We got two plasmids from the Fussenegger group at the ETH Zurich containing a codon optimized version of bpsA (pMM64) and svp (pMM65), respectively, for expression in HEK 297-..."</p>
<p><b>New page</b></p><div><br />
We got two plasmids from the Fussenegger group at the ETH Zurich containing a codon optimized version of bpsA <br />
<br />
(pMM64) and svp (pMM65), respectively, for expression in HEK 297-3 cells [Muller 2012]. The first goal is to <br />
<br />
transform the plasmids into competent E. coli (Rosetta) and check functionality of the plasmids in our cells.<br />
<br />
===Indigoidine production with pMM-plasmids I (Ilia)===<br />
After Transformation cells are grown first on Amp medium and thereafter on Kan medium.<br />
* transform competent Rosetta with 225 ng pMM065 and 253.5 ng pMM064<br />
* plate on Amp plate<br />
* pick colonies from Amp plate, make liquid cultures in LB + Kan + IPTG(1 mM)<br />
* Evening: no growth in liquid culture => prepare ON culture in LB+Amp<br />
* Put ON culture in incubator<br />
* [[Heat-shock competent bacteria|prepare competent]] Rosetta-pMM064 from ON culture<br />
* transform with 225 ng pMM065<br />
* plate on Kan+IPTG plate<br />
* no growth on Kan plate<br />
<br />
===Results and Discussion===<br />
There were no blue colonies. Transformation will be repeated and medium will be provided with both antibiotics.</div>FloSmi