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Q5 High Fidelity DNA Polymerase

 

Q5 High-Fidelity DNA Polymerase sets a new standard for both high fidelity and robust performance. The Q5 buffer system is designed to provide superior performance with minimal optimization across a broad range of amplicons regardless of GC content.

"I am impressed with the PCR efficiency and accuracy of Q5!"

-Marjorie Mercier

Marjorie Mercier, a scientist in Brussels, shares her positive experience with Q5 High-Fidelity DNA Polymerase for the amplification of the IgM variable region.

We needed a polymerase that could handle templates with a varying GC content including GC rich. Microdessected germinal center cells were used to clone the V(D)J combination in IgM for expression in cells. Therefore we need the correct sequence to keep the ORF intact. We don’t want to introduce errors by the polymerase.

We performed a nested PCR on IgM transcripts yielding a PCR product from about 500-600 bp with variable GC content. We tested the new Q5 High Fidelity polymerase against Phusion High Fidelity polymerase. The results clearly show a much better yield for Q5 then Phusion.

The fragments have been completely sequenced and were devoid of any mistake.

 

Reaction Set-up

Component 25 µl Reaction Final Concentration
5x Q5 Reaction Buffer 10 µl 1x
5 mM dNTP 1 µl each 200 µM each
10 µM Forward primer (1st or 2nd) 1 µl

0.5 µM

10 µM Reverse primer (1st or 2nd) 1 µl

0.5 µM

Template DNA (cDNA or 1st PCR product 1/100)

2.5 µl

< 50 ng

Q5 High-Fidelity DNA Polymerase

0.25 µl

0.5 U

Nuclease-Free Water

To 25 µl

To 25 µl

 

PCR Protocol

Step Temperature Time

Initial Denaturation

98 °C

30 seconds

20 / 25 Cycles

98 °C

10 seconds

65 °C / 70 °C

30 seconds

72 °C

15 seconds (+1 sec / cycle)

Final extension

72 °C

5 minutes

Hold

10 °C

0.5 U

Ordering Information

Cat. # Description Quantity

M0491S/L/XL

Q5™ High-Fidelity DNA Polymerase

100/500/2500 units

M0492S/L

Q5™ High-Fidelity 2X Master Mix

100/500 rxns

M0493S/L/XL

Q5™ Hot Start High-Fidelity DNA Polymerase

100/500/2500 units

M0494S/L

Q5™ Hot Start High-Fidelity 2X Master Mix

100/500 rxns

E0555S/L

Q5™ High-Fidelity PCR Kit

50/250 rxns

B9027S

Q5™ Reaction Buffer Pack

6 ml

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