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Third round PCR

PCR Molecular Biology

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#1 matongo

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Posted 07 November 2012 - 03:54 AM

Hi!
I ran a classical first and second round PCR reaction. I got a very weak band of my desired product from the 2nd round PCR. After many trials, I dicided to run a third PCR round from the product and conditions of the 2nd round; I finally got a big band of the desired product. But after purification with ZYMOGEN, I still experience a negative value of the ratio 260/230 indicating a contamination. So is the problem with the 3th PCR round or the cleaning up prossess? My product is about 1Kb.
Thanks

#2 bob1

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Posted 07 November 2012 - 11:38 AM

Definitely the cleaning process - pure DNA doesn't really read at the 230 nm range, but phenolics, proteins, and quite a few other things do.

#3 matongo

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Posted 12 November 2012 - 05:34 AM

Thanks





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