2 replies to this topic

[PCR Novice]

Hi

In our department here we have been setting up a PCR lab, for over a year now, and have run into a major problem with cDNA contamination-it just seemed to keep spreading and spreading no matter what we did, I have been reading numerous blogs here and getting tips and I think we finally have it under control.

The problem I have is that we prepared a Maxi-prep and I seem to have contaminated it with cDNA-we do what we call an 'elegant negative' where we do a PCR and use primers that shoudnt work on the plasmid, it worked a month ago but with the amount of contamination we had we wanted to repeat it before we took the next step. Is there anyway I can clean-up the plasmid without having to redo the Maxi-prep???

Thanks

[molgen]

Try running your plasmid on a gel.
Extract it from the gel and then redo the PCR.

[phage434]

Given the sensitivity of PCR, I think it is unlikely that a gel purificaton will remove template contamination. Perhaps it is worth a try.