I am planning to do SNP genotyping using Allelic Discrimination technique. I am going to use Applied Biosystems 7500 Fast Real-Time PCR system. I saw so many Taqman master mixes on their websites. Since I am not an expert in molecular techniques and this is my first experience, I want to order the proper master mix for doing this technique. Could anybody clarify for me the difference between universal master mix and genotyping master mix?
TaqMan Genotyping Master Mix is optimized for end-point fluorescence detection in SNP genotyping applications. TaqMan Universal Master Mix is an improved master mix formulation for real-time PCR. It has been validated for gene expression, SNP genotyping, and microRNA research
ABI recommends using Genotyping Master Mix for SNP allelic discrimination. Is is formulated to increase specificity and discrimination. They do not offer the Universal mastermix for TaqMan assays anymore. The Gene expression mastermix is optimized for yield.
If you have primers and probes take any master mix for TaqMan from ABI, BioRad or other for system evaluation. If you using ABI allelic discrimination sets - use what ABI (ThermoFisher) reccomend
ABI has good plateform for genotyping, and if you doing 5 allele discrimination then needs universal master mix other wise ABI recommends using genotyping Master mix for SNP allelic discrimination.
Please find enclosed file.
All the best
TaqMan Genotyping Master Mix is optimized for end-point fluorescence detection in SNP genotyping applications. TaqMan Universal Master Mix is an improved master mix formulation for real-time PCR. It has been validated for gene expression, SNP genotyping, and microRNA research
TaqMan Universal Master Mix (with uracil-DNA glycosilase which prevents carryover contamination from previous amplicons has a green label in the box, otherwise is red) it is considered the gold standard for real time PCR in ABI thermocycylers because it can be applied to genotyping, gene expression, etc. It is versatile (cheaper if yoo are planning to do more than one application) but not optimized for each application. On the other hand TaqMan genotyping master mix is an optimized mix for genotyping that is suitable for facilicities where robots are available (as 7500 HT) because it present higher stability during more hours at room temperature, e.g. overnight or weekend runs. Also, in my experience, this master mix solves difficult genotype calls (due to GC- or AT-rich regions) that TaqMan Universal Master Mix can not.
Thank you all for informative answers.
Mr Ernesto; I was also wondering about the difference between master mixes with AmpErase® UNG and without? Could presence of AmpErase® UNG improve Allelic Discrimination? I am going to use 7500 Fast Real Time PCR system from Applied Biosystems for this technique.
I appreciate your advice. Thank you.
It is better with AmpErase due to you will have more confidence that your genotype is correctly assigned and there is no remaining amplicons from previous runs. Also be careful and avoid to contaminate the plate/tube with your own skin cells (no uncover skin between your lab coat and gloves).
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