User:Kathryn Muratore/Notebook/AU CHEM-570 lab prep/2011/07/06

AU CHEM-570 Lab Prep

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Objective Improve transformation efficiency Bench work Transform NovaBlue 2.5μL V(His)+I ligation from last week + 25μL NovaBlue 2.5μL V(His) neg ctrl ligation from last week + 25μL NovaBlue follow transformation protocol heat shock is 80s @ 42°C add 450μL SOC for recovery → plate 100μL on LBAmp100 → 37°C O/N Re-plate transformants spread 450μL of DH10B transformants from yesterday on LBAmp100 → 37°C O/N Electrocompetent DH10B - starter culture 3 mL LB + 50μL E. coli DH10B from thawed chemically-competent cells → 37°C O/N w/ shaking Culture ER2566 for glycerol stock and also to confirm that media is okay since there have been problems with some of my students growing starter cultures. 3 mL LB + colony E. coli ER2566 from streaked plate → 37°C O/N w/ shaking Test Amp100 #* 3 mL LB + DH10B/pKSeAATHis colony #1 from one of Tamra's transformation tests this week x 2 → 37°C O/N w/ shaking Results No growth on yesterday's plates