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I am preparing competent cells, and I finished inoculating a single colony in SOB. It has been incubating at 37 degrees Celsius for almost 16 hours since, and it's getting very late where I live. I wanted to do the large-scale inoculation a few (at least 7) hours from now instead of right away, but I do not want the cells to start dying. Can I take the cells out of the incubator and leave the cells in room temperature? I'm hesitant about this because even at room temperature, wouldn't cells still be growing, making it possible for cells to die as well? What extra steps, if any, should I take before using the cells?

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    $\begingroup$ Don't do this. User fresh cells thoughout the process, everything else can affect your transformation efficiency in the end. $\endgroup$
    – Chris
    Jun 10, 2015 at 7:35
  • $\begingroup$ What would you suggest? $\endgroup$
    – wswr
    Jun 10, 2015 at 7:40
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    $\begingroup$ I always did the preculture (picking a single colony from the plate) overnight (12-16 hrs typically) and then started the main culture from which I harvest the cells for the competent cells the next morning. Here you need to carefully check the OD anyways to not overgrow the cells. $\endgroup$
    – Chris
    Jun 10, 2015 at 7:41
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    $\begingroup$ I would use a fresh overnight culture then. A colleague tested this a while ago and the cells which are stored for a while give less efficient transformation rates. $\endgroup$
    – Chris
    Jun 10, 2015 at 7:55
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    $\begingroup$ What kind of inoculation? What is the final purpose? It may not be a problem at all. Ask someone in your lab to take the cells out and keep them in 4⁰C $\endgroup$
    – WYSIWYG
    Jun 10, 2015 at 8:22

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