Methods for primer reconstitution - (May/03/2006 )
Hello all,
I ordered primers from Operon.They recommend reconstituting primers in TE buffer at pH 7.0.Can somebody help me out on this.What concentrations of Tris and EDTA should I use to prepare TE and can I use pH other than 7.0 (7.0-8.0 maybe?).They r not recommending reconsitution of primers in sterile,distileed water.Does that mean I can't reconstitute primers in nuclease free water as well?
did you search on the various topics on that in the forum?
http://www.protocol-online.org/forums/inde...wtopic=7660&hl=
http://www.protocol-online.org/forums/inde...wtopic=7790&hl=
I ordered primers from Operon.They recommend reconstituting primers in TE buffer at pH 7.0.Can somebody help me out on this.What concentrations of Tris and EDTA should I use to prepare TE and can I use pH other than 7.0 (7.0-8.0 maybe?).They r not recommending reconsitution of primers in sterile,distileed water.Does that mean I can't reconstitute primers in nuclease free water as well?
tris 10 mM EDTA 1 mM pH 7.0-7.5 maybe 8.0 is OK. nuclease free of course.
in sterile water it will be less stable and more difficult to resuspend if pH<7.0 (and it is often the case with deionized water).
Fred I will take care of this in the future.
Missele many thanX for ur help!!
well, i want to precise that this was just a question and not an attak 
i just did the search for you and post the relevant topic.
I did not say it with wrong intentions Fred...I should have searched for the relevant topic 
On a side note I am thinking of reconstituting my primers in nuclease free water as I don't think ti's a good idea to prepare TE buffer manually becoz for that I shall need millipore purified water or distilled water and I don't think it will be available in my lab.I might end up introducing contamination in my primers. I am not in the position of taking any risk.What do u ppl suggest?
use the nuclease free water to prepare the te and use that.
Not possible!!
sure it is
you treat with DEPC
then autoclave(twice to be sure)
then add nuclease-free tris and edta (you can get it from sigma, mol biol grade) with your best technique
then autoclave again
you r making me even more confused! 