Third round PCR - (Nov/07/2012 )
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.
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.