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PCR, neg control & no. of cycles

pcr neg control contamination

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2 replies to this topic

#1 ImNew

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Posted 19 October 2011 - 02:41 AM

Hi guys,

In my previous thread, I stated that I started to use 50 cycles for my PCR.

After gel electrophoresis, I realize that there is a single band that is of the same size as my desired PCR product in
my negative control.

This is likely due to contamination of the reaction mixture with DNA template, right?

It shouldn't have anything with the process itself (50 cycles of PCR), right?

Thanks.

Regards,
ImNew

#2 Papaver

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Posted 20 October 2011 - 10:45 PM

It's strange that only your negative control shows a band. If you used a mastermix I would expect that you would see a band in ervery reaction.

#3 phage434

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Posted 21 October 2011 - 04:43 AM

50 cycles of pcr is absurd. Nothing happens after about 40. Likely your template has an inhibitor which is preventing the amplification of your contaminating DNA. You could try diluting your template 10x to 100x.





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