PCR amplification using REDTaq® ReadyMix™ PCR Reaction Mix

November 03, 2014

BlueskyReddit
Newcastle University
Academic Haematology
Research Associate

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Company:

Sigma Aldrich

Product Name:

REDTaq® ReadyMix™ PCR Reaction Mix

Catalog Number:

R2523

See Similar Products:  PCR Master Mixes

I have used the REDTaq ReadyMix PCR reaction mix to amplify mutation containing regions of DNA for downstream Sanger sequencing.

Experimental Design and Results Summary

Application

Amplification of DNA regions for Sanger sequencing

Starting Material

DNA extracted from tumour samples

Protocol Overview

The protocol is very straightforward. Briefly, a reaction mix is prepared based on 50ul volume, comprising 25ul ReadyMix, typically 0.5uM forward and reverse primer, 10ng template DNA and water. The components are mixed and amplified using a thermal cycler under standard conditions.

Tips

All reagents are contained within the ReadyMix for ease of use.

Results Summary

Results indicate strong amplification, although it should be noted that this Taq is reported to be less stringent than competative reagents. However, it is suitable for the validation applications such as confirming known mutations by Sanger sequencing.

Additional Notes

The red dye enables quick visual confirmation of reaction mixing. PCR products can be loaded directly onto an agarose gel following PCR.

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Summary

The Good

Rapid and simple protocol for PCR with all amplification components in one mix.

The Bad

Less stringent than other Taqs

The Bottom Line

Economical and simple to use product that can be loaded straight into gels without the need for loading dye.

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