HiFi PCR Mix for NGS

Item Number

H665865

• Storage Temp: Store at -20°C,Avoid repeated freezing and thawing
• Shipped In: Ice chest + Ice pads

Product Description

Product content

H665865	Component	1 mL	5 mL	Storage
H665865A	2×HiFi PCR Mix	1 mL	5×1 mL	-20℃. Avoid freeze/thaw cycle.
H665865B	ddH₂O	1 mL	5×1 mL	-20℃. Avoid freeze/thaw cycle.

Product Introduction

HiFi PCR Mix for NGS is a premixed system consisting of hot starter enzyme, PCR Buffer, dNTPs, Mg2+, and PCR stabilizers and enhancers, etc. It is characterized by high fidelity, high elongation, and low preference, and has a balanced amplification efficiency for complex DNA templates (e.g., high GC-content templates), and it is especially suitable for multiplexed PCR in the thousand-two library building .

The high efficiency hot starter enzyme contained in this product has no polymerase activity at room temperature, which effectively avoids non-specific amplification caused by non-specific binding of primers and templates or primer dimerization under room temperature conditions. The unique combination of buffer system and hot starter enzyme significantly improves the amplification efficiency of PCR with wider amplification range. This product effectively enhances the amplification efficiency of high GC or high AT regions in the genome, reduces amplification preference and thus improves sequencing coverage.

matters needing attention

This product is not suitable for thousands of primers with modifications.

Before use, please mix gently by turning up and down, avoid foaming as much as possible, and use after brief centrifugation. Avoid repeated freezing and thawing, which may deteriorate the performance of the product. If frequent use is required within a short period of time, store at 2-8°C.

Usage

The following examples are conventional PCR reaction systems and reaction conditions, which should be improved and optimized according to the template, primer structure and target fragment size in actual operation.

PCR reaction system

Reagents	50 μl Reaction system
2xHiFi PCR Mix	25 µI
Primer Pool	0.1-0.3 µM
DNA or cfDNA	5 ng-100 ng
ddH2O	up to 50 µl

Note: Please use the final concentration of 0.1-0.3µM as a reference for setting the range of primer concentration.

2. PCR reaction conditions

Attention:

In general, the annealing temperature was 5°C lower than the melting temperature Tm of the amplification primers, and when the desired amplification efficiency could not be obtained, the annealing temperature was lowered appropriately; when non-specific reactions occurred, the annealing temperature was increased, and the reaction conditions were optimized.

2) The number of cycles can be set according to the downstream application of the amplification product. If the number of cycles is too low, the amplification is insufficient; if the number of cycles is too high, the chance of mismatch will increase and the non-specific background will be serious. Therefore, the number of cycles should be minimized under the premise of ensuring the product yield.