Master Mix (PCR)

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Below is the recipe for making a 2x Master Mix stock solution for use in PCR reactions. Ensure that each reagent is thoroughly thawed and pipette the full volume of each before starting to ensure that they are thoroughly mixed.

Recipe

The below table shows the recipe for approximately 10 reaction mixtures. Some reagents may be substituted for other manufacturers/catalog numbers, especially for Nuclease Free Water, but I included these values for what I used with mine. For the Lai lab, all of these reagents can be found in the center -20ºC freezer at the end of Blaine's bench in a box called "PCR Pre-mix" with an orange tape label.

Note

Typical master mixes for PCR make a 1X solution, but the water amount has been halved here to make a 2x solution. This is so that when your master mix is added to your sample later on during experiments and it gets diluted, it will approach 1X. The below protocol is for a master mix solution of 500 μL.

Reagent Manufacturer Cat# Amount
Nuclease Free Water Technova W3330 340 μL
10X PCR Buffer ( - MgCl2), DNA-free† Invitrogen y02028 100 μL
50 mM MgCl2, DNA-free† Invitrogen y02016 30 μL
10 mM dNTP mix‡ Denville CB4421-6 20 μL
Platinum Taq DNA Polymerase, DNA-free (5 U/μL) Invitrogen 100021274 10 μL
Total 500 μL

†Indicates reagents that should be included with the Platinum Taq DNA Polymerase kit.

‡Indicates that the reagent is made from a stock solution that must be diluted to reach the concentration shown.