2 x Fast Pfus PCR Master Mix (for PCR series)
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หมวดหมู่ : Biochemicals , 1. Chemical and Reagents , Buffers , Servicebio ,
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Product Information
Product Name | Cat. No. | Spec. |
2 x Fast Pfus PCR Master Mix | G3305-01 | 1 mL |
G3305-05 | 5×1 mL |
Product Description/Introduction
This Fast Pfus PCR Master Mix is a ready-to-use PCR premix containing modified Pfu DNA polymerase, dNTPs and optimized PCR reaction buffer at a concentration of 2× for routine PCR, colony PCR, PCR with complex templates and high GC templates. PCR amplification can be performed by adding only the template, primers and ddH2O so that the concentration of Mix solution is 1×. Strong amplification ability, fast amplification speed, extension speed up to 5-15 s/kb, high fidelity, high specificity, amplification product is flat end. Loading Dye has been added to the product, and the amplified product can be directly used for agarose gel electrophoresis detection.
Storage and Shipping Conditions
Ship with wet ice; Store at -20, valid for 12 months. Avoid repeated freeze-thawing.
Product Contents
Component | G3305-01 | G3305-05 |
2 x Fast Pfus PCR Master Mix | 1 mL | 5×1 mL |
Manual | One copy | |
Assay Protocol / Procedures
1. Recommended PCR reaction system
Component | 20 μL rxn | 50 μL rxn | Final Concentration |
Templatea | Variable | Variable | as required |
Forward Primer (10 μM)b | 0.8 μL | 2 μL | 0.4 μM |
Reverse Primer (10 μM)b | 0.8 μL | 2 μL | 0.4 μM |
2×Fast Pfus PCR Master Mix | 10 μL | 25 μL | 1× |
(DMSO,optional)c | (0.6 μL) | (1.5 μL) | (3%) |
ddH2O | Add to 20 μL | Add to 50 μL |
a: Higher amounts of template increase the risk of generation of non-specific PCR products. Lower amounts of template reduce the accuracy of the amplification. If the template is plasmid or from bacteriophage, 50ng-5pg added in 50 μL rnx is recommended. If tempalted is genomic DNA, 250ng-50ng added in 50 μL rnx is recommended. If template is cDNA, dilute your original cDNA by 2-100 times, and the volume of diluted cDNA added to reaction is no more than 10% of total reaction volume. If templtae is bacterial fluid or crude extract sample, the volume added to reaction is no more than 10% of total reaction volume. Higher amounts of template tend to lead to non-specific amplification. Lower amounts of template tend to lead to low PCR amplification efficiency.
b: The range of final primer concentration used is 0.2-1.0 μM，and 0.4 μM is recommended. Too little primer may result in low or no amplification yield, and too much primer may result in non-specific amplification.
c: For high GC content template, DMSO with no more than 10% total volume.
2. Recommended PCR amplification conditions
Step | Temperature | Time | Number of Cycles |
Initial Denaturationa | 98 | 30 s-120 s | 1 |
Denaturation | 98 | 5-10 s | 25-35 |
Annealingb | 50-72 | 10-30 s | |
Extensionc | 72 | 5-15 s/kb | |
Final extension | 72 | 5-10 min | 1 |
Hold | 4-16 | Forever |
a: A pre-denaturation time of 30 s is suitable for most conventional templates; complex templates can be denatured for up to 2 min.
b: For amplification of complex templates, containing concatenated primers, the annealing time can be extended up to 30 s.
c: The recommended extension step is 5-10 s/kb at 72° for simple templates such as plasmid ,10-15 s/kb for routine genomic DNA templates, 15-30 s/kb for complex template.
Note
1. When using, please thaw and mix thoroughly, after completely thawed can be placed in 4 stable storage at least 2 weeks, avoid repeated freezing and thawing.
2. For your safty and health,please wear safety glasses, gloves, or protective clothing.
For Research Use Only!
