2×A8 FastHiFi PCR MasterMix (with dye)
Lot # Check on the product label
Size: 5 ml
Storage and Expiration: Store at -20°C for 2 years.
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Price: $140.00
Catalog# BPC1019-2
Lot # Check on the product label
Size: 5 ml
Storage and Expiration: Store at -20°C for 2 years.
Description: This product contains A8 FastHiFi DNA Polymerase, dNTPs and optimized reaction buffer at a concentration of 2×. When using, just add template, primer, and make up water to the final concentration of Mix to 1×. A8 is derived from genetically engineered pfu, which greatly improves the amplification speed, fidelity and yield. A8 Mix is the first choice for ultra-fidelity and rapid amplification of non-long fragments. This product contains a red tracer dye. It can be directly loaded for electrophoresis without adding a loading buffer; it can also be purified for subsequent operations such as restriction digestion, ligation, and fluorescence sequencing.
Recommended PCR system settings
|
Components |
25 µl reaction |
50 µl reaction |
Final Concentration |
|
2× A8 PCR MasterMix |
12.5 μl |
25 μl |
1 × |
|
Forward Primer(10 μM) |
0.5 μl |
1 µl |
0.2 μM |
|
Reverse Primer(10 μM) |
0.5 μl |
1 μl |
0.2 μM |
|
Template DNA |
as required |
as required |
|
|
ddH2O |
up to 25 μl |
up to 50 μl |
|
Reference template dosage (50 μl reaction system):
Plasmid: 0.1-10 ng; bacterial genome: 10-100 ng; human genome: 50-150 ng; cDNA: 1-5 μl from RT reaction.
Recommended PCR cycling conditions
|
Step |
Temperature |
Time |
Cycle Number |
|
Initial denaturation |
95°C |
3 min |
|
|
Denaturation |
95°C |
10 s |
30 cycles |
|
Annealing |
55°C |
10-15 s |
|
|
Extension |
72°C |
15-30 s/ kb |
|
|
Final Extension |
72°C |
2-5 min |
|
|
|
4-8°C |
Hold |
|
Precautions
- The amplification speed of A8 is fast. Simple templates such as plasmids and simple genomes can be used at 15-20 s/kb; complex templates such as human genomes can be used at 30-45 s/kb.
- For templates with high GC content, the pre-denaturation and denaturation temperature can be increased to 98°C. A8 has strong heat resistance, 98°C does not change the activity of A8.
- If the GC content of the amplified template is high or the template is complex and the amplification effect is not good, can add DMSO to the reaction mixture to a final concentration of 1%-8%, and follow a 1% gradient increase to find the optimal concentration. Or add betaine to a final concentration of 1.0-1.7 M. And use Touchdown PCR.
