* The sequences have been double-checked
Has anyone had a PCR stop working after changing primer manufacturers? They might use different purification techniques. The amount of lyophilized primer might not be accurate on the label leading to bad dilutions. My PCR works with one brand but not the other. Ideas why?
It is not necessary unless there is a good reason for doing so. One good reason could be poor synthesis of one primer so assuming that the syntheses are the same scale you expect to dissolve the primers in the same amount of TE to get the same concentration so if your stock original primers were in 600ul and one new primer is in 600ul but the other needs dissolving in 200ul then the amount of this last primer is very suspicious and something has gone wrong in the synthesis and it needs to be replaced. I never found any significant difference between desalted oligos and FPLC/HPLC/gel purified oligos in pcr so would not suspect the purification method when troubleshooting pcr
Thank you! It's good to know that I don't need to worry overmuch about the purification. Perhaps my next step should be checking how the working stock is being prepared. You make a very good point: the SOP might give a volume rather than a concentration for resuspension. I'll have to take a look.
Cheers! Very helpful.
Not necessary, although how you prepare your working stock may be look into, to fall inline with the new manufacturers guidelines and dilution ratios.
Thank you! Yes, taking a look at their recommendations is a good idea. :)
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