Haynes Lab:Notebook/Characterizing AHL quorum sensing homologs/2013/10/21

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mm/dd/yyyy

Retry ligation from 10/19/13 again with gel extracted J06702
Backbone: J06702 E/X: 44 ng/ul

Cut, gel purified

Insert: Gblck pLac-RBS E/S: 2.6ng/ul

Cut, column purified

same calculations from before, still only actually tried 3:1 and bb ctrl

1:1 2:1 3:1 Bb ctrl
insert 0.5 1.0 1.5 0
vector 1.13 1.13 1.13 1.13
2x Roche ligation buffer 5 5 5 5
T4 ligase 1 1 1 1
H20 2.37 1.87 1.37 2.87


ligations ran for 3 hours at RT
Incubate with 50ul DH5αT for 45 min on ice
heat shock for 30s at 42°C
ice for 2min
spread



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