I am doing qPCR in Corbett machine and having some problems with a really high efficiency. Iíve tried almost everything that Iíve read could cause that problem:
1. Increase the Temperature
2. Change the amount of the MgCl2, which was a slightly better, but still 130-150 % efficiency
3. Change the amount of the primers
4. Played with the dNTPs
5. Make different dilutions
By now I am using only DNA from Promega (female) and this is the only one thing that I didnít change just because it is standard DNA and I donít think it could be the problem. Have anyone had the same problems with the standard DNA? And do you have any ideas what could couse the problem?