This kit from Fermentas includes Nuclease-Free Water and 2X PCR Master Mix: a 2X concentrated solution of
Taq DNA Polymerase, dNTPs and all the components required for PCR. All the user provides is the template and template-specific primers. The pre-mixed formulation saves time and reduces contamination due to a reduced number of pipetting steps required for PCR set up. Its applications include high throughput PCR, routine PCR, generation of PCR products for TA cloning (generates PCR products with 3'-dA overhangs), and RT-PCR. The 2X PCR Master Mix is sufficient to perform 100 standard 50 µl PCR reactions (with 1.25 units of
Taq DNA Polymerase).
I use this kit for general PCR amplification as well as for production of product for use with Invitrogen’s TOPO TA Cloning Kit, in which the PCR product is ligated into a thymidine-overhang vector for sequencing. The pieces I am sequencing are very small, approximately 50-70 base pairs. This 2X master mix seems to work well for this purpose and my sequencing success rate seems to be around what it was when I was making master mix myself; it has neither positively or negatively affected my sequencing results.
What I like most about the kit is the pipetting it saves, although in reality it doesn’t save that much, just the few steps that go into making master mix. Each kit contains around 2.5 ml of master mix and I use them up like mad - my typical 50 µl reaction uses 5 µl of sample, 2.5 µl of each primer, 15 µl water and 25 µl master mix. Initially, I would aliquot the master mix into 0.65 ml tubes, but this proved to be a waste of time (and master mix) and I would use all the aliquots in one go.
The bottom line is, I am on the fence with this one. I am not sure that it saves me much time or money, but I will continue using it for the time being because it does save me a few pipetting steps.
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Shelly D. Roper
Chemistry Technician
Air Force Research Labs, HEPC
Conceptual Mindworks, Inc
United States
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