Haynes Lab:Notebook/Characterizing AHL quorum sensing homologs/2014/05/12

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mm/dd/yyyy Ran PCR products from 5/6/14 on a gel. Only 6, RhlI worked. Re-ran other PCRs, annealing temp 40, 4min ext Number Template F Primer R Primer Annealing T Expected length 1 pTet-mCh 2/14/14 74ng P127 P097 53 3148 2 LuxI sender const 8/31/13 179ng P063 P064 23 654 3 K084007 3/10/13 4-2-1 P069 P070 32 645 4 K084007 3/10/13 4-2-2 P069 P070 32 645 5 RBS-LasI 3/8/13 31ng P069 P070 32 645 Ran a second one of the backbone with annealing temp 50, ext time 4 min Label Template F Primer R Primer Annealing T Expected length mCh sender pTet-mCh 2/14/14 74ng P127 P097 53 3148 Ran on gel, only 5, LasI worked. Ran a third, annealing temp 30 Label Template F Primer R Primer Annealing T Expected length LuxI LuxI sender const 8/31/13 179ng P063 P064 23 654 Realized P127 and P097 will never work on pTet-mCh because after the first round with one primer, the binding site for the other one is at the wrong end. Will need to redesign one of the primers to add a second RBS in front of mCherry or behind pTet.