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Cell Maintenance Media with Selection Agents (Zeocin, Blasticidin, Geneticin)
F12 from Gibco, 10% FBS, 1% Penicillin/Streptomycin, 1/10,000 Blasticidin, 1/4,000 Zeocin, 1/500 Geneticin
Blasticidin = 10mg/ml stock (pcDNA6)
Zeocin = 100mg/ml stock (pcDNA4)
Geneticin, G418, stock (pcDNA3, pIRES)
- Obtain a 50mL conical tube and label as:
- “Ham’s F-12 Media + 10% FBS + 1% Pen/Strep + 1ug/ml blast + 25ug/ml zeo + 100ug/ul gen”
- Include date made and initials
- Add 44.5ml of Ham’s F-12 Media to the labeled 50mL conical tube
- Add 5ml of FBS
- Add 500µl of Pen Strep
- Add 5µl of blasticidin (Final Concentration (1µg/ml))
- Add 12.5µl of zeocin (Final Concentration (25µg/ml))
- Add 100µl of geneticin (Final Concentration (100µg/ml))
- Mix by pipetting
- Close the conical tube tightly and store in 4°C
To make selection media for establishing new cell lines, use 10x more selection agent for final concentrations of:
- Add 50µl of blasticidin to 50ml of Ham’s F-12 Media + 10% FBS + 1% Pen/Strep media (Final Concentration (10µg/ml))
- Add 125µl of zeocin to 50ml of Ham’s F-12 Media + 10% FBS + 1% Pen/Strep media (Final Concentration (250µg/ml))
- Add 1000µl of geneticin to 50ml of Ham’s F-12 Media + 10% FBS + 1% Pen/Strep media (Final Concentration (1000µg/ml))
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