I have a temperature-sensitive plasmid that gets cured at 42 degrees and simultaneously expresses a protein under the regulation of a temperature-sensitive lambda repressor.
For its curing and protein expression in E.coli cells, it should be put in a shaking water bath and allowed to grow for 20 minutes. The rapid temperature shift is important for protein expression, so it is suggested to use a shaking water bath. I do not have access to a shaking water bath. Are there any suggestions as to how I can make sure about the temperature shift?
Also, are there ways to check whether the plasmid was cured and the protein was expressed?
If the protein gets expressed, I get colonies on an antibiotic plate.
Dear Shilpi Kumari
which is the volume scale of your cultures?
Are you sure that
Since for small volumes you can use also different shaking platfrom as eppendorf Thermomixer, since if the sample volume is small the temeprature shift will be enough fast also in this kind of platform.
Differently is if you would like to perform high volumes. In this case the best way is probably a fermenter however also if i'm not expert of this kind of induction system, i'm quite optimist that performing cultures with a low ratio culture volume/shaking flask volume and buffled cup vented shaking flask for which air exchange is faster, your sistem can work also in a standard orbital shaker.
good luck
Manuele
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