I had a confusion regarding tag buffers when I was running my PCR reaction and I accidentally pulled out a vial of MgSO4 as opposed to MgCl2. My PCR ran blank and I am kind of wondering if it has to do with the salts. Since I was not using any of the high fidelity tags like pfu or anything I am curious to know if MgSO4 is still not a good choice of buffer with normal taq polymerase. Has anyone experienced similar problems when using the sulfate buffer?
Standard Taq polymerase should work okay with magnesium sulfate.
In theory no, many Taq PCR buffers contain ammonium sulfate (instead of KCl). Varying the amount of Mg2+ ions will affect your PCR however - I suggest you repeat the PCR, using the right vial!
Hi
No problem. Many commercial Pfu + Taq DNA polymerases are supplied with magnesium sulfate.
God speed you
my research experience suggest that use of MgSO4 or MgCl2 donot have any effect as many a times I have used interchanged. it is the concentration 5that matters. again in theory also Mg++ is required for enzyme activity. therefore I advice you to repeat the experiment again.
Abbas, Cl2 or SO4 are interchangeable for the vast majority of Taq polymerases out there. In fact I haven't found one that will not work well with MgSO4. The possible issue is the concentration of the MgSO4 vs the MgCl2 you use. Taq is fairly robust across a broad range of input Mg++ but it can be inhibited if the Mg++ conc is too high (>5mM) or too low (80% activity) is usually between 1.5 and 2.5 mM Mg++ for various Taq preps in my experience. Higher Mg++ is more inhibitory in my hands, but it seldom causes complete shutdown of activity.
Check the conc of the SO4 vial you used and see how much Mg++ you actually added to your PCR reaction.
I assume that there was no EGTA running around your nucleic acid prep?
- Badges
- Science method