AQ97 Hot Start High Fidelity DNA Polymerase (AQ97 HS HiFi) is a thermostable, chimeric DNA polymerase created specifically for low-bias, high fidelity amplification of a vast range of amplicons. AQ97 HS HiFi delivers high-speed elongation and processivity, due to its fusion with a DNA-binding domain.
The antibody-mediated inactivation of the polymerase and exonuclease functions increases specificity of the enzyme, as well as enabling room temperature setup.
AQ97 HS HiFi is recommended for applications, which require extremely high fidelity such as cloning/sub-cloning, NGS applications, SNP analysis and mutagenesis.
FEATURES
2 minute hot start
Reaction setup at room temperature
High fidelity: >60x Taq Fidelity
High elongation rate: 10 sec/kb
3’→5’ proofreading exonuclease activity
Long range capacity: 18 kb for gDNA and 25 kb for λ DNA
Good coverage on DNA templates with low to high GC content
DESCRIPTION
AQ97 HS HiFi exhibits both 5'→3' DNA polymerase activity and 3'→5' proofreading exonuclease activity, enabling accurate correction of base-pair mismatches. The attached DNA-binding domain enhances processivity, while the antibody-mediated inactivation ensures specificity even at room temperature setup. Together, these features yield an enzyme that consistently delivers high-fidelity, specific, and fast results with long-range capacity and low-bias amplification.
The AQ97 HS HiFi kit includes the 5x AQ97 Buffer allowing robust amplification of long targets with low to high GC content. The kit also contains an extra tube of MgCl2 in 25 mM concentration if adjustments are required.
Addition of 1-2 M Betaine Enhancer Solutions is recommended when amplifying difficult amplicons, such as GC-rich targets, long targets, or those with complex secondary structures.
For more convenient handling AQ97 HS HiFi is also available as a 2x master mix.
FIDELITY
The fidelity of a polymerase refers to its ability to insert the correct base during PCR. On the contrary, the rate of misincorporation is known as a polymerase error rate. Fidelity depends on the polymerase, the buffer system in use and the quality of your template DNA.
Performance of AQ97 Hot Start High Fidelity DNA polymerase (AQ97 HS HiFi) was compared to three leading hot start high fidelity DNA Polymerase (Q,S and P). Eight different human genomic DNA targets, 400 – 800 bp in length and with GC content ranging from 29 – 78 %, were amplified. Robust amplification was observed for all targets using AQ97 HS HiFi. DNA Polymerase Q provides very similar results to AQ97 HS HiFi. The same is the case for DNA polymerase S, except for the target with 78% GC content, where AQ97 HS HiFi performs better. In contrary, DNA polymerase P was not able to provide the same level of robust and specific amplification of DNA targets >70%, under the conditions tested here. Amplification studies has been set up, as recommended by the manufactures. Tm calculators of the respective competitors were used to calculate optimal annealing temperatures for primers. When amplifying GC-rich targets, 2 M Betaine Enhancer Solution (AQ97 HS HiFi), GC enhancer (DNA Polymerases Q and S) or GC-rich specific PCR Buffer (DNA Polymerase P) was included in the reaction mix. Marker M: Iqon PCR Ladder from Ampliqon (A610641). Click to enlarge picture.
LONG-RANGE AMPLIFICATION
Performance of AQ97 Hot Start High Fidelity DNA (AQ97 HS HiFi) polymerase on large and complex amplicons was compared to three leading hot start high fidelity DNA Polymerase (Q, S and P). Six different targets of human genomic DNA ranging from 2 kb to 17.5 kb were amplified. Robust amplification was observed for all targets using AQ97 HS HiFi. DNA polymerases Q, S and P generally yielded higher PCR product quantities, except for the last and longest target, where both DNA polymerases Q and S resulted in weaker bands and less PCR product. DNA polymerases Q, S and P also tend to produce more background noise and nonspecific bands, particularly noticeable in the first and fourth targets. Marker M: High Range DNA Ladder from Ampliqon (A610141). Click to enlarge picture.
