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PCR without DNA extraction!


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#1 sarigsk

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Posted 25 May 2009 - 04:49 AM

Hi

Is there any way to perform PCR on genomic DNA from eukaryotic cells without the need to extract DNA (e.g., for checking if a cell contains a specific construct after transfection)?

Any advice will be highly appreciated
Thanks in advance

Edited by sarigsk, 25 May 2009 - 05:44 AM.


#2 tea-test

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Posted 25 May 2009 - 06:04 AM

there is something from finnzymes but i never tried it
http://www.finnzymes...l?Ad=homeDirect
tea-test: The artist formerly known as Ned Land

#3 warsel

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Posted 28 May 2009 - 04:45 AM

Hi

Is there any way to perform PCR on genomic DNA from eukaryotic cells without the need to extract DNA (e.g., for checking if a cell contains a specific construct after transfection)?

Any advice will be highly appreciated
Thanks in advance


There are ways, for instance just taking your cells of interest in some TE, boiling for 15min and then using this in a PCR can work.
The problem for your application is, however, that your medium will still contain plasmid from the transfection which you might not be able to wash away very well.
You should, therefore, take some medium along in a PCR as well as a negative control; otherwise your experiment doesn't tell you much.

I usually check for transfection efficiency by co-transfecting some GFP plasmid with the plasmid of interest. That way I always know if my transfection was good or not on the next day.




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