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which G418 concentration and treatment period should be used?


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#1 feifei83

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Posted 23 March 2011 - 10:12 AM

I need G418 to select transfection cells. I treat cells before transfection with G418 in 6-well plates to determine the proper G418 concentration (the test concentration is from 100-1100ug/ml, and G148 was used when planting cells.)
Then I found that, for my cells,
1, after 4D treatment, cells in g418 1000ug/ml all died.
2, after 6D treatment, cells in g418 700ug/ml all died.
3, after 7D treatment, cell in G418 500ug/ml all died.
4, after 8D, cells in G418 400ug/ml all died.
5, after 9D, cells in G418 300ug/ml all died.
(For cells in wells with G418 100-500ug/ml, I added new medium with the same concentration of G418 7 days after first 7D treatment.)
So, my problem is which G418 concentration should I choose? And how long should I treat my cells for selection?
Thanks.

#2 bob1

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Posted 23 March 2011 - 02:39 PM

Use the minimum needed to kill untransfected cells after the recommended treatment time (IIRC for G418 = 7-10 days), so somewhere between 330-500 ug/ml.

#3 feifei83

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Posted 24 March 2011 - 06:21 AM

thanks. try to use 400ug/ml first.




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