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Taqman qPCR protocols are run on a real time PCR instrument. Use an appropriate instrument specific Fluorophore/Quencher combination.
Mutant= 85 bp
Wild Type = 119 bp
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
40114 | GCT TTC AAG TTT CAT CCA TGT TG | Common | A | mChr17 | ||
40115 | TTC ATG ATA GAC AGA TGA ACA CAG T | Wild type Reverse | A | mChr17 | ||
40116 | Fluorophore-1 | TGT GTG TCA AGA GTG GTT TTT TAT TCT A | Quencher-1 | WT Probe | ||
40117 | GTC TTT GGG TAA AGC CAA GC | Mutant Reverse | A | Mt1 | ||
40118 | Fluorophore-2 | TGT CAA GAG TAC CAA GCT TAT GGG | Quencher-2 | MUT Probe |
Component | Final Concentration |
---|---|
Kapa Probe Fast QPCR | 1.00 X |
ddH2O | |
40114 | 0.40 uM |
40115 | 0.40 uM |
40117 | 0.40 uM |
Wt Probe | 0.15 uM |
Mutant Probe | 0.15 uM |
DNA |
Step | Temp °C | Time | Note |
---|---|---|---|
1 | 95.0 | -- | |
2 | 95.0 | -- | |
3 | 60.0 | -- | |
4 | -- | repeat steps 2-3 for 40 cycles | |
5 | 40.0 | -- | Forever |