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Hot Start Taq DNA Polymerase
Description
The Hot Start Taq DNA Polymerase is a recombinant Taq DNA polymerase which has been chemically modified by the addition of heat-labile blocking groups to its amino acid residues. The enzyme is inactive at room temperature, avoiding extension of non-specifically annealed primers or primer dimers and providing higher specificity of DNA amplification. The functional activity of the enzyme is activated in 10 minute incubation at 95°C. The activated enzyme maintains the same functionality as Taq DNA polymerase: Has the 5′to 3′DNA polymerase activity and 5′to 3′exonuclease activity without 3′-5′exonuclease activity. The extending speed is 1 kb/30 sec. There is an "A" on 3′end.The PCR product can be cloned in TA vector.
Applications:
Hot-start PCR amplification
Specific amplification of complex cDNA and genomic template
Amplification from low copy number DNA template
Real-Time PCR
Multiple PCR
Generation of PCR products for TA cloning
Quality Control:
Functional absence of double and single-stranded endonuclease activity; Purity>99% test by SDS gel electrophoresis; Each lot of HotStar Taq DNA Polymerase is assayed for amplification from as little as 10 ng of human genomic DNA ; Retain full activity at room temperature for one week; No host DNA residue.
Concentration: 5 u/μl
Storage: Store at -20°C
Reaction Mixture Set Up
Component Volume Final Concentration
Template DNA <1 ug as required
Forward Primer (10 μM) 2 μl 0.4 μM
Reverse Primer (10 μM) 2 μl 0.4 μM
10×HotStarTaq PCR Buffer 5 μl 1×
2.5mM each dNTPmix 4 μl 200μM each
HotStar Taq DNA polymerase, 5U/μl 1μl 2.5 unit
ddH2O to final volume 50μl Not applicable
Note: the supplied buffer is 10X and contains 15mM Mg2+,although 1.5mM Mg2+ works well for most amplification, the optimal [Mg2+] is determined empirically. Be aware of any chelators in samples.
Recommended thermal cycling conditions
94℃ 10 min
94℃ 30 sec
55-65℃ 30 sec 30-35 cycles
72℃ 30 sec
72℃ 5-10 min
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