primer contamination - primer contaminated with ice (Nov/20/2009 )

Hi all,

I am glad to have found this site. I apologize if this topic has already been discussed else where but I searched and could not find a specific answer to my problem.

When I began my mastermix on ice, a small piece of ice accidentally fell into one of the primers. I tried to remove the ice but couldn't and decided to continue with the the PCR reaction anyway to see what the results would be. Before the ice totally melted however, I used the automatic pipette to withdraw some primer from the bottom portion of the tube. Is the primer ruined for good and how will ice effect PCR reaction?

Please help. Any input is appreciated!

Thanks.

I don't think the presence of ice itself would cause a problem (I've always made up my reactions on ice or in a cooled container), but if you just get your ice from a shared ice machine (like I always have) then you've probably contaminated your primer - the effect is probably dependent on what you're targeting. If you're amplifying algal specific genes, you might be ok, but if you're looking at something more ubiquitous...anyway its probably a good idea to replace your primer. Sorry and good luck!

mairimike on Nov 20 2009, 12:50 PM said:

Unfortunately I did get my ice from a shared ice machine. I will go ahead and replace my primer and redo. Thanks for your quick response Shannon.

I agree with Shannon. While you may be lucky and whatever contamination you get from the ice does not affect your experiment, the fact of the matter is that the last thing you want to do is introduce any uncertainty into your experiment, especially from something as inexpensive as a primer.

When I resuspend my primers I always do it on a benchtop, at room temperature, and away from everything else. Nucleic acids resuspend very well at room temperature and in the ~5 minutes that it takes to resuspend the primers you will not degrade.