potassium acetate in primer solution - I made a mess but need to recover (Feb/09/2006 )

I just made a mistake, I was making primer 10uM solution but I used protein preciptation solution
instead of water by mistake.
I am not a moleculer biology person so I need to ask you that;
this primer cannot be recovered?
what will be happen if I use this primer solution for PCR reaction. pottassium acetate will
affect the reaction or what will be expected?
I hope someone can help me.

Hi

You should be able to recover the primer by precipitating it with ice cold isopropanol and precipitating it in the freezer for 1-2 hours. pellet the DNA in a microfuge at max revs and resuspend in a small volume of water or TLE. Don't forget to quantify.

BTW for primers most people make stocks of 100 uM and dilute them down to 10 uM for working with, this avoids having big volumes of the initial solution.

I would guess that having the acetate in the reaction will affect the PCR if you attempted to amplify the DNA, mostly this would be due to inhibition of the reaction by a pH change, though if you used anough primer you could possibly precipitate the taq, not that you would want to.

Cheers,
Bob

hi
i agree with bob1.
Do a isopropanol precipitation and wash 2-3 times with 70%ethanol in order to remove salts (which is not permitted with isopropanol.

Bob and Fred, thank you so much for your help.
I forgot to tell that I made 50uM stock solutions first and 10uM working solutions (both with Potassium acetate) so I have 50uM ones to perform precipitation process.
I am going to try that.
Daisy

I just want to say thank you for you guys.
recovery rates were 75-90% for my 3 different primers. (I used Nanodrop to measure the conc. of DNA)
I tried PCR runs with therse primers and got good results from all of them.

Daisy

your welcome.