Optimizing PCR reactions for gene amplification – Master Mix

OVERVIEW

If you are working in the geomicrobiology field or a geologist (or just curious) )who is interested in geomicro you will run into PCRs (Polymerase Chain Reaction) at some point in time. Basically the PCR lets you target a piece of DNA and then copies it a billion times or so. This means lots of it! Once you have this mess of segments you can then identify what it is. Pretty neat. The University of Utah has an excellent Virtual PCR program. I’d recommend checking it out.

WHY?

So you collected some rocks that have buggies on them. You might want to know who the buggies are. Or maybe you are interested in seeing if they have genes that produce siderphores (compounds the scavenge metals). PCR can help with this (so can 454, which is another topic).

We use AmpliTAQ Gold because we can get it downstairs fairly cheaply. Not all TAQs are created equal. I am going to run an experiment later with different TAQs to compare each one.

Basic Master Mix recipe for AmpliTAQ gold

for 15 µL reaction

(volumes in µL)

H2O 9.57

10x PCR Buffer 1.5

100x BSA 0.125

Forward Primer 0.1

Reverse Primer 0.1

TAQ 0.15

MgCl2 1.5

There you go.

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