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opugirl 12-07-2012 10:17 PM

Annealing primers
 
So I'm doing a relatively simple ligation process

The purpose is to construct a vector by
1-digest Vector A w/ YYYI and gel purify
2-anneal primers containing ZZZI/XXXI site and
3-then ligate the oligos with the digested Vector A fragment

My problem is I did this years ago and a coworker let me borrow a reagent they used for annealing the primers what I termed the annealing buffer. It was from an "Expanded Template PCR System" I remember I had to heat it up to dissolve all this white stuff in the bottom and that it was very viscous. I'm guessing it was some sort of salt. Maybe any salt would work (MgCl2?)

Anneal Primer Mix
4 ul annealing buffer
2ul forward primer from 10mM oligo stock
2ul Reverse primer from 10mM oligo stock
32ul dH2O

I have the program for the thermocycler that worked very well last time so that is all set. Any suggestions as to what this salt buffer could be or what could work would be super super appreciated! I have access to most majorly used PCR reagents from life technologies, promega, etc.


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