2X Universal Taq Master Mix

2X Universal Taq Master Mix

Description:
Universal Taq Master Mix (2X) combines high-quality MCLAB recombinant Taq DNA Polymerase, a recombinant hot start protein, and MCLAB Ultrapure nucleotides in a proprietary reaction buffer. This ready­to-use mix provides robust and reliable performance for demanding PCR applications in which high specificity and high sensitivity are desired. Since the mix is pre-formulated, experimental variability is significantly reduced. It would be used for PCR amplification up to 8 kb .
Applications:
High-specificity PCR amplification High-sensitivity PCR amplification TA cloning
High throughput PCR
Convenient and Usage:
The pre-mixed formulation saves time and reduces potential contamination and eliminates pipetting errors. For a 20 µl reaction, simply add 10 µl of 2x Universal Taq Master Mix to primers, DNA template and PCR­Qualified H2O. Reactions can be easily performed in 10 µl, 25 µl, 50 µl or 100 µl volumes. Room temperature reaction assembly is possible because of the hot start feature.
2x Universal Taq Master Mix Formulation
2x Universal Taq Master Mix combines MCLAB proprietary Taq DNA Polymerase in a unique buffer formulation. Magnesium and nucleotide concentrations are 7.0 mM and 0.4 mM each, respectively.
Storage:
Shipped on dry ice. Store at -20°C. Mix well prior to use.

Brief Protocol
This standard protocol applies to a single reaction where only template, primers, and water need to be added to the master mix. For multiple reactions, scale-up volume of reaction components proportionally.

1. Thaw reagents at room temperature. Mix thoroughly and then place on ice.
2. Assemble reactions on ice or at room temperature, whichever is more convenient.
3. The following table shows recommended component volumes

   Components	Vol. for 20 µl reaction	Final Concentration
   Universal Taq Master Mix (2X)	10 µl	1 x
   10 pM Forward Primer	1  µl	0.2 pM
   10 pM Reverse Primer	1  µl	0.2 pM
   Template DNA	1-100 ng for genomic DNA or plasmid DNA	As needed
   Water, PCR-Qualified	up to 20 µl	N/A

1. Ensure reactions are mixed thoroughly by gentle vortexing followed by a brief spin in a microcentrifuge.
2. The following table shows recommended cycling conditions:

   Cycle Step	Temperature	Time
   Initial Denature	94-95°C	2 minutes
   Repeat following three cycles as necessary, generally 25- 35 times.*
   Denature	94-95°C	30 seconds
   Anneal *	55°C	30 seconds
   Extend **	72°C	60 seconds per 1 kb
   Final Extend	72°C	5 minutes
   Final hold	4°C	as necessary

* 45 cycles may be required for low-copy targets.
** Initially, annealing temperature should be 5°C below the calculated Tm of the primers. If non-specific products are produced, increase the annealing temperature in 1-2°C increments.
*** Extension time should be about one minute for every kilobase of expected product size.
6. Analyze sample (typically 1 to 10 pl aliquots) by agarose gel electrophoresis. Visualize PCR product in gel with DNA intercalating dyes and an imaging device.