Master mix for multiplex PCR
For general laboratory use.
Shipping: shipped on gel packs
Storage Conditions: store at -20 °C
avoid freeze/thaw cycles
Shelf Life: 12 months
Concentration: 2x conc.
Multiplex PCR Master is specially designed for the set-up of multiplex PCR reactions. It contains an optimized composition of polymerase, nucleotides, MgCl2 and stabilizing components in a specifically developed buffer system allowing the parallel amplification of a multitude of fragments in a single PCR assay.
The master mix contains all reagents (except primer and template) in a 2x concentrated ready-to-use solution.
The kit is recommended for use in routine PCR reactions and highly suitable for multiple target gene amplification in a single tube.
The high specificity and sensitivity of the mix is achieved by a chemically inhibited hot-start polymerase. Its activity is blocked at ambient temperature preventing the extension of nonspecifically annealed primers and primer-dimer formations at low temperatures during PCR setup.
2x Multiplex PCR Master (red cap)
master mix containing Hot Start Taq polymerase, nucleotides, optimized reaction buffer and stabilizers
PCR grade water (white cap)
Recommended 50 μl PCR assay:
Prepare a master mix of all components except template to reduce pipetting errors. A reaction volume of 20-50 μl per assay is recommended for most PCR cyclers. Pipet with sterile filter tips and perform the setup in an area separate from DNA preparation or analysis. No-template controls should be included in all amplifications.
|component||stock conc.||final conc.||50 μl|
|Multiplex PCR Master||2x||1x||25 μl|
|forward primer 1||10 μM||400 nM||2 μl|
|reverse primer 1||10 μM||400 nM||2 μl|
|forward primer 2||10 μM||400 nM||2 μl|
|reverse primer 2||10 μM||400 nM||2 μl|
|forward primer ...||10 μM||400 nM||2 μl|
|reverse primer ...||10 μM||400 nM||2 μl|
c) plasmid and lambda DNA
a) 10-200 ng
b) 1 - 50 ng
c) 1 - 5 ng
|PCR-grade water||-||-||fill up to|
Recommended cycling conditions:
|95 °C||12 min||1x|
|Denaturation||95 °C||30 sec||30 - 50x2)|
|Annealing1)||58 - 64 °C||40 sec||30 - 50x2)|
|Elongation3)||72 °C||1 min/kb||30 - 50x2)|
|72 °C||5 min||1x|
3)The elongation time depends on the length of the fragments to be amplified. A time of 1 min/kb is recommended.
For optimal specificity and amplification an individual optimization of the recommended parameters may be necessary for each new template DNA and/or primer pair.
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