Haynes Lab:Notebook/Characterizing AHL quorum sensing homologs/2014/05/21

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mm/dd/yyyy

Golden gate 1:10 RhlR and EsaR receivers grew but there were colonies on the backbone plate

Ligation Colonies
Bb 13
RhlR 23
EsaR 14



Ran PCR, extension time 3:30 at 50°C

Template F primer R primer
pTet-mCh 130 97
LuxI GFP rec 131 132
LuxR 1:10 colony 1 103 104
LuxR 1:10 colony 2 103 104
LuxR 1:10 colony 3 103 104
LuxR 1:10 colony 4 103 104
LuxR 1:10 colony 5 103 104
LuxR 1:10 colony 5 103 107


Ran above PCR on gel

Template F primer R primer Expected band size Actual band size
pTet-mCh 130 97 3130 3130
LuxI GFP rec 131 132 610 no band
LuxR 1:10 colony 1 103 114 1817 no band
LuxR 1:10 colony 2 103 114 1817 1817
LuxR 1:10 colony 3 103 114 1817 no band
LuxR 1:10 colony 4 103 114 1817 no band
LuxR 1:10 colony 5 103 114 1817 1817
LuxR 1:10 colony 5 103 107 1035 1035



picked LuxI and RhlI GFP senders into 4mLs media and shook overnight to screen for successful receiver transformations.