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No growth following transformation

PCR Transformation Culture

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#1 kmmargart

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Posted 18 July 2012 - 02:31 PM

I have been attempting to transform our competent cells and have hit a stumbling block. Along with my sample I set up two controls; one pUC18 control, and one mutagenic control. The problem is that after transformation and streaking my plates, the only growth I have is from the pUC18 control with a 99.33% mutagenesis efficiency. I repeated the transformation on a new batch of competent cells hoping for something different but ultimately ended with the same results. I have tried to find an answer but am coming up empty handed, I'm an undergrad student participating in some research and could use any advice. Could there be an issue during the PCR process that would cause this, or could it be related to the types of cells being used? Please help!

Thanks,
Kenny

#2 pcrman

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Posted 20 July 2012 - 05:04 PM

Could you describe in more detail what you have done? You mentioned PCR, what is mutagenic control...

So you obtained colonies from an intact control plasmid, but not from your mutagenic control. Is that caused by problems in ligation or other things so that the construct did not confer your competent cells antibiotic resistance?





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