User:Daniel Goodman/Notebook/Cluzel/2010/04/05

Cluzel Lab Notebook Daniel Goodman

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Prepping Plasmid for Electroporation PZE-12-CFP vector from JM Kim Vector Nomenclature: http://www.expressys.com/main_vectors.html Electroporations ng/ul 260/280 260/230 56.3 ng 1.92 1.62 Doing 4 electroporations, in order to ensure competent cells made last week work correctly. Symbol Media Plasmid Cells Why A LA/Amp PZE-12-CFP My competent MG1655 To get comp cells w/ plasmid B LA PZE-12-CFP My competent MG1655 To check that my competent cells are alive C Spec/AMP PZS4-ATTP My competent MG1655 To check that my competent cells transform D LA/AMP PZE-12-CFP Jeff's competent MG1655 To check that my plasmid works/is correct Protocol steps on page 1.119 - 1.122 of Molecular Cloning, Volume 1 book used straight glycerol instead of GYT used SOC medium (1 ml per reaction) 1 ul of DNA per experiment (approx 52 ng for my plasmid, did not nanodrop Jeff's spec plasmid) 1.8 kV, 1 second shock Put in 37 degree shaker w/ the SOC for 1 hour. Spun down, supernatant removed to ~50ul each, resuspended, plated and spun onto correct antibiotic plates w/ LB. Put in 37 degree incubator overnight at 7:45 pm. Will take out tomorrow morning.