Product Information

Product Description

      This product contains Profusion™ DNA Polymerase, dNTPs and optimized reaction buffer at a concentration of 2×. When using, just add template, primers, and add water to the final concentration of Mix to 1×. Profusion™ DNA Polymerase is a mixture of a variety of ultra-high-fidelity enzymes modified by the most advanced genetic engineering with extension factors added , which greatly improves the amplification length, amplification speed, fidelity and yield. Profusion™ PCR Master Mix is the preferred product for ultra-fidelity and rapid amplification of long fragments. This product contains red tracking dye and can be directly loaded for electrophoresis without adding loading buffer; it can also be purified for subsequent operations such as enzyme digestion, ligation, and fluorescent sequencing. The PCR product has blunt ends and does not need to add A heads. It can be directly cloned using the Holly pTOPO Blunt blunt-end series vectors (Cat. No. HRF0161 , HRF0171 ).

      2×Profusion™ PCR Master Mix has the advantages of being fast, simple, highly sensitive, highly specific, and stable. The reaction system only requires the addition of primers and templates, and can be amplified through a two-step procedure, simplifying the experimental steps and saving time.

Features

1. Fast amplification speed: The extension speed can reach 3-4kb/min, which is 6-8 times that of pfu.
2. High amplification yield: The amount of PCR product is generally 50%-100% higher than the traditional pfu yield.
3. Excellent fidelity: The fidelity is more than 54 times that of taq. Generally, a random bacterial sequencing is the correct colony without mutation.
4. Long amplification length: Using complex genomic DNA as a template is suitable for amplifying products of no more than about 10kb, and using simple genomes, plasmids and phage DNA as templates is suitable for amplifying products of no more than about 15kb.

Application

Gene cloning; amplification of complex DNA templates; high-throughput library construction.

Transportation and storage methods

Transported with ice packs. Store at -20℃, valid for 2 years.

Precautions

1) Crystals may precipitate in the Master Mix after thawing. Be sure to vortex and mix until the precipitate disappears before use to avoid affecting the performance of the reagent.

2) For your safety and health, please wear a lab coat and disposable gloves when operating;

3) This product is for scientific research purposes only!

Recommended PCR reaction system (prepared on ice)

Reference template dosage (50 μl reaction system):

Recommended PCR amplification procedure

Notes:
1. 2×Profusion™ PCR Master Mix has a fast amplification speed. Simple templates such as plasmids and simple genomes can be amplified at 10-15 seconds/kb; complex templates such as the human genome can be amplified at 20-25 seconds/kb.
2. If the main band is found to be blurred up and down in electrophoresis, or there is a smear band dragged down from the sample well in the lane, it generally indicates that the extension time is too long. The gradient shortens the extension time (10-15 seconds/kb is recommended as the gradient to reduce the extension time) until satisfactory results are obtained.
3. For templates with high GC content, the pre-denaturation and denaturation temperatures can be increased to 98°C. Profusion™ DNA Polymerase has strong heat resistance, and 98°C has no effect on the activity of the enzyme.
4. If the amplification template has a high GC content or the template is complex and the amplification effect is poor, DMSO can be added to the reaction mixture to a final concentration of 1%-8%, and the optimal concentration can be found by increasing the gradient by 1%. Or add betaine to a final concentration of 1.0-1.7M. And use touch down PCR (Touch down PCR)