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Download Catalog 2009/2010

HotRox Master Mix 2x

PCR-Mastermix information : The number of reactions is calculated on 50 µl per reaction.
For reseach use only.

We offer our Mix in two different MgCl2 concentrations. Specifically for TaqMan PCR were higher MgCl₂ concentations were recommended the mix with 7,0 mM MgCl2 makes optimization much easier.

Datasheet: HotRox Master Mix 2x

Description:

2 x HotRox Master Mix is an optimized ready-to-use PCR mixture of Taq DNA Polymerase, antibodies to Taq DNA Polymerase, PCR buffer, MgCl2 and dNTPs. 2x PCR Master Mix contains all components for PCR, except DNA template and primers. ROX Reference dye is included in the HotRox Master Mix to normalize the fluorescent signal on instruments that are compartible with this option. The mixture was shown to be effective for Real Time PCR.

2x HotRox Master Mix Composition:

• Taq DNA Polymerase (recombinant) in reaction buffer: 0.1 unit/µl 
• Antibodies to Taq DNA Polymerase, concentration adjusted for the effective inhibition of DNA polymerase activity at 37^oC     
• 32 mM (NH₄)₂SO₄
• 130 mM TrisHCl, pH 8.8 at 25^oC  
• 0.02% Tween-20  
• 3 mM MgCl₂  /7 mM
• dNTPs (dATP, dCTP, dGTP, dTTP): 0.4 mM of each
• ROX dye 1µM/0,1µM

Instrument	Final ROX Conc.
ABI 7000, 7300, 7700, 7900HT	500 nM
ABI 7500, Stratagene Mx3005P, Mx4000	50nM

Pack size:

2x 1,25 ml - sufficient for 100 HotStart PCR reactions in 50 µl reaction volume

Performance and purity tests :
Tested for the absence of endodeoxyribonucleases and exodeoxyribonucleases. The 2x HotRox Master Mix is tested in the amplification of a single-copy gene of mouse genomic DNA.

Endodeoxyribonuclease Assay:
No detectable conversion of covalently closed circular DNA to nicked DNA was observed after incubation of 25 µl of 2x HotRox Master Mix with 1 µg of pUC19 DNA in 50 µl for 4 hours neither at 37°C nor at 70^oC.

Application: 
- PCR
- Primer extension
- Multiplex PCR
- Low-copy targets PCR  
- Real-time PCR  

Associated activities: 
Endonuclease and exonuclease activities were not detectable after 2 hours incubation of the mixture with 0.22 mg of EcoR I digested lambda DNA at 72^oC in the presence of 15-20 units of enzyme in HotRox Master MIX  

Storage conditions:
Store HotRox Master MIX frozen at -20°C and protect from light. 

Protocol for PCR with SuperHot PCR Master Mix

Due to the inhibition of polymerase activity at room temperature by Anti Taq DNA polymerase antibodies all reactions may be settled-up at room temperature, it will not result in increase of unspecific product or primer-dimers formation.

Add in a thin walled PCR tube:

	50 µl reaction volume		25 µl reaction volume
component	volume	final conc.	volume	final conc.
2x PCR Master Mix	25 µl	1x	12.5 µl	1x
Forward Primer	variable	0.1-1 µl	variable	0.1-1 µl
Reverse Primer	variable	0.1-1 µl	variable	0.1-1 µl
Template DNA	variable	10 pg-1µg	variable	10 pg-1 µg
Sterile Deionized Water	up to 50 µl	-	up to 25 µl	-

• Gently vortex the sample and briefly centrifuge to collect all drops to the bottom of the tube.
• Overlay the sample with mineral oil or add an appropriate amount of wax if the thermal cycler is not equipped with a heated lid.
• Place the samples in a thermocycler and start the optimal PCR program.      

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