This product is a pre prepared mixture of all reagents, dNTP, and PCR enhancers required for genotyping PCR reaction, with a concentration of 2 times.
2 × Genotyping Taq PCR MasterMix is designed and optimized for rapid amplification detection of conventional PCR and complex templates such as genomic DNA. This product is convenient and fast to use. Simply add primers and templates and dilute with deionized water to 1 × You can proceed with the reaction.
2 × Genotyping Taq PCR MasterMix offers two forms for you to choose from: regular type and rapid sample loading type. After testing, the addition of dyes does not affect amplification performance. After the PCR reaction is completed, electrophoresis can be performed directly, saving time.
This product is suitable for amplifying trace template DNA from complex backgrounds. Due to the higher mutation rate of Genotyping PCR compared to regular PCR reactions, its products are not recommended for gene cloning that requires high fidelity.
The primers used in Genotyping PCR need to be carefully designed, otherwise more heterobands will be amplified (see the golden rule of PCR primer design).
Application examples:
Comparison of amplification sensitivity detection: 2 × Taq PCR MasterMix (left image) VS 2 × Genotyping Taq PCR MasterMix (right image)
Using a concentration of 100ng/ μ The concentration of human genomic DNA will be 1ng/ μ Dilute the plasmid containing LacZ gene gradient to the following concentration gradient: 0.1ng/ μ l. 0.01ng/ μ l. 1pg/ μ l. 0.1pg/ μ l. 0.01pg/ μ l. 1fg/ μ Take 1 from each concentration gradient μ As an amplification template, use 2 × Taq PCR MasterMix and 2 × Genotyping Taq PCR MasterMix amplifies the LacZ gene (each undergoes 40 rounds of PCR amplification cycles).
Result display: 2 × Genotyping Taq PCR MasterMix detection sensitivity ratio 2 × Taq PCR MasterMix is over 1000 times higher. Swim lane M: DNA Ladder 2000; Swimming lanes 1-6: corresponding to the concentration gradients of each template above
Common reaction systems:
Component volume
two × Action Mix * 10 μ L
Upstream primer 0.05uM (final concentration)
Downstream primer 0.05uM (final concentration)
Template DNA 1-3ul DNA sample
Taq enzyme (5U/ μ l) 0.25 μ L
DdH2O to 20 μ L
*The reaction system can be proportionally adjusted to 50 μ L Equal volume
Common PCR cycles:
93 ℃ for 1 minute and 30 seconds
40 cycles at 93 ℃ for 30 seconds
57 ℃, 30 seconds (57 ℃ is suitable for most reactions and can also be adjusted between 55-60 ℃)
65 ℃, calculated based on product length of 60 seconds/1KB
4 ℃ insulation
