I apologize for the very naive question, but I'm just starting out in a high school biology lab and I am very confused. If I have stocks of 100μM for forward and reverse primers separately, I can dilute them to 10μM very easily. But what if I want to make a master mix? I have 1000μL of 10μM for each primer, forward and reverse (not mixed). Would I just add 500μL of my forward primer and 500μL of my reverse primer for a mix of 1000μL in which both concentrations are still 10μM?
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Would I just add 500μL of my forward primer and 500μL of my reverse primer for a mix of 1000μL in which both concentrations are still 10μM?
No. Diluting your 10μM solution in half will half the concentration. Mixing equal parts of 10μM primer will make a master mix where each primer is 5μM
But in general, primers are added to these reactions in great excess, so 5μM of primer might be okay.
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$\begingroup$ The question also states that they have 100 uM stocks of each primer. In that case, they can make their master mix (with 10 uM of each primer) using those stocks. $\endgroup$ Commented Mar 1, 2019 at 20:49