The first few cycles of PCR are always the most important to successful amplification. Our mastermixes are never short of enough polymerase to kick start and then powerfully continue your amplification. MegaMix Gold uses a superior sensitive hot start DNA polymerase. The polymerase becomes active upon heating at 95C. This ensures a highly specific and sensitive amplification and ensure no background or primer dimer formation.
Using Microzone’s own proprietary engineered PCR buffers, our mastermixes provide powerful, broad template amplification. PCR products generated are A-tailed and suitable for ligating into TA cloning vectors.
- When you need Hot-Start you can rely on Gold.
- Exceptionally stable and can be freeze thawed on multiple occasions.
- High success rates up to 6kb.
- High yields under standard conditions.
- Provides effective and efficient amplification for GC and AT rich templates.
- Strong MgCl2 to support strong amplification.
- Microzone PCR Buffer – provides increased stability, performance and ease of use.
- HotStart Routine end point PCR
- qPCR – needs additional dye to be added
- Fast PCR (not multiplexing)
- Multiplex PCR
- Amplification of GC and AT rich templates
- Up to 6kb
- TA Cloning
- More confidence in amplification and PCR test
- Mastermix format (Ready to use or 2x) for ease of use
- Utilises Microzone’s own unique PCR buffer offering enhanced PCR and stability.
- Easy set up and PCR optimisation
- Broad range of templates and conditions
Thaw the MegaMix
Add the desired volume (25µl in a 50 µl reaction, 10µl in a 20µl reaction etc.) into the PCR tube.
Add gDNA Template – 10 – 200ng. Less is always more.
cDNA Template <100ng
- Final forward and reverse concentration of 0.2 – 0.6 µM from a 10 µM stock
Overlay with mineral oil if necessary
- Place in a Thermal Cycler
Cycling Profile (guide only)
- Initial denaturation step: 95⁰C for 3 mins
- Then Cycle 25-30 times:
- Step 1: 95⁰C for 30 secs
- Step 2: Optimal annealing temp. of primers for 30 to 60 secs
- Step 3: 72⁰C for 45 to 60 secs
- Cool to room temperature
- Thaw reagents prior to use. Mix well prior to use.
- GC and complex templates may require higher denaturation temperatures
- Two step cycling protocols can increase speed. Following annealing just ramp the temperature back to denaturation.
- If using new primers or changing your mastermix it may be necessary to perform a new annealing temperature gradient. We suggest 1⁰C intervals and to start 5⁰C below the calculated Tm of the primers and go to 5⁰C above the primer Tm.
- Primers design and quality have a large impact on the reaction. Taking time to design your primers using good techniques or software will benefit down the line. We do offer a technical note on primer design. Contact us to receive a copy.
MegaMix Gold is available to be purchased with an organisation’s purchase order. For payment by debit / credit card, please visit our UK distributor Clent Life Science.