Difference between revisions of "PCR 2X Master Mix"
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:0.5 µl 10mM dNTPs | :0.5 µl 10mM dNTPs | ||
:1 µl 50mM MgCl<sub>2</sub> (adjusting the Mg concentration affects the specificity of the polymerase and can be optimized) | :1 µl 50mM MgCl<sub>2</sub> (adjusting the Mg concentration affects the specificity of the polymerase and can be optimized) | ||
| − | :0.5 µl (5 U / µl) Taq polymerase | + | :0.5 µl (5 U / µl) thermostable (Taq) polymerase |
:8 µl nuclease-free H<sub>2</sub>O (adjust if necessary if MgCl<sub>2</sub> or Taq volume is adjusted for a 12.5 µl total volume). | :8 µl nuclease-free H<sub>2</sub>O (adjust if necessary if MgCl<sub>2</sub> or Taq volume is adjusted for a 12.5 µl total volume). | ||
In general a mix would be made for many reactions, say for 100, and the volumes multiplied accordingly. The mix can be stored at -20 C. | In general a mix would be made for many reactions, say for 100, and the volumes multiplied accordingly. The mix can be stored at -20 C. | ||
Revision as of 16:15, 16 July 2018
The master mix can be purchased premixed commercially or mixed from the individual components.
For a single 25 µl reaction
- 2.5 µl 10X buffer (usually this is supplied with the polymerase, this assumes it does not already have
- 0.5 µl 10mM dNTPs
- 1 µl 50mM MgCl2 (adjusting the Mg concentration affects the specificity of the polymerase and can be optimized)
- 0.5 µl (5 U / µl) thermostable (Taq) polymerase
- 8 µl nuclease-free H2O (adjust if necessary if MgCl2 or Taq volume is adjusted for a 12.5 µl total volume).
In general a mix would be made for many reactions, say for 100, and the volumes multiplied accordingly. The mix can be stored at -20 C.
