Haynes Lab:Notebook/Engineering PC-TFs/2013/09/07: Difference between revisions
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== | ==September 7, 2013== | ||
* | '''Grow liquid culture of CMV promoter''' | ||
*Label 15 ml sterile culture tubes. Fill each tube with 2 ml LB growth medium with Amp. | |||
*Use sterile pipette tip to touch bacterial streak and put the tip into LB medium. | |||
*Grow the cultures for 10-12 hrs. | |||
[[Image:cmv.jpg|200px|CMV promoter]] | |||
'''Miniprep liquid cultures''' | |||
*Add 100 μL of 7X Lysis Buffer (Blue) to 600 μL of E.coli culture in a 1.5 ml microcentrifuge tube. Mix by inverting tube 4-6times. | |||
*Add 350 μL of cold Neutralization Buffer (Yellow), mix thoroughly. Neutralization is complete when sample becomes yellow and precipitate has formed. | |||
*Centrifuge at 11,000-16,000 g for two minutes. | |||
*Transfer the supernatent into the Zymo-Spin Column. Place column in 2 ml collection tube and centrifuge for 15 seconds at top speed. Discard flow-through. | |||
*Add 200μL of Endo-Wash Buffer to the column. Centrifuge for 15 seconds at top speed. | |||
*Add 400μL of Zyppy Wash Buffer to the column. Centrifuge at top speed for 30 seconds. | |||
*Transfer column to 1.5 microcentrifuge tube and add 50μL of Zyppy Elution Buffer directly to column. Centrifuge for 1 minute at top speed to elute DNA. | |||
Latest revision as of 23:17, 26 September 2017
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September 7, 2013Grow liquid culture of CMV promoter
Miniprep liquid cultures
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