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Large quantity heat shock trouble?

transformation E. coli heat shock

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3 replies to this topic

#1 jungskwon98

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Posted 14 December 2012 - 07:41 AM

Hi I'm working in a lab where we need to large quantities of transformation

Using heatshock method and E. Coli.

10min on ice , 30 sec heat shock , 2 min on ice plate

When I plate my ligation mixture using single tube for transformation they work fine.

But for large quantity(96 well pcr plate) if the transformations work they give very low yield

or don't work at all. Is there any advice anybody can give me?

Thank you

#2 phage434

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Posted 14 December 2012 - 06:33 PM

My guess is that your heat shock is insufficient in the plate you are using. There is a wide range of plate format. What kind of plate are you using? A deep well plate, for example, will behave very differently than a pcr plate. I'd suggest doubling your heat shock time to see if it has a positive effect.

#3 Trof

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Posted 15 December 2012 - 02:07 AM

First run heat-shock duration optimisation on the type of well you are using. One by one, I think it's worth if you want to do 96 at one time later.

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#4 jungskwon98

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Posted 17 December 2012 - 08:59 AM

Thank you all for the reply I will first optimize my temp and time. Will update you later





Also tagged with one or more of these keywords: transformation, E. coli, heat shock

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