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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 = 89 bp
chr15:78191610-78191698 89bp GCAGAATTCTCCACTCTGGTG GACTGAGATGGGGCGTTG
Wt Sequence:
Mutant Sequence:
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
50343 | TCT AAT TCC ATC AGA AGC TGG T | Mutant Forward | A | |||
50344 | GAC TGA GAT GGG GCG TTG | Common | A | |||
50345 | GCA GAA TTC TCC ACT CTG GTG | Wild type Forward | A | |||
50346 | Fluorophore-1 | AAA AGC TTC TGC TTG GGT CCC C | Quencher-1 | MUT Probe | ||
50347 | Fluorophore-2 | AAG TGG CGC TGA CTG CTT GG | Quencher-2 | WT Probe |
Component | Final Concentration |
---|---|
Kapa Probe Fast QPCR | 1.00 X |
ddH2O | |
50343 | 0.40 uM |
50344 | 0.40 uM |
50345 | 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 |