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Taqman qPCR protocols are run on a real time PCR instrument. Use an appropriate instrument specific Fluorophore/Quencher combination.
Mut= 99 bp
Wt= 100 bp
Fam=Mut
Hex=Wt
Wt Sequence (deletions in lower case:
AAGGCAGTTTTTGCCTCCTTGTGGCTGGCGGTCACACTAACGGATGGCCCTGCATACTTTGTGtttgacgcaggttctgggctgacaaacatcaagacggaagagatctcggaagtgaagatggatgcagaattcggacatgattcaggatttgAAGTCCGCCATCAAAAACTGGTAGGCAAAAATAAAC
91bp del
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
40445 | AAG GCA GTT TTT GCC TCC TT | Common | A | |||
40446 | CCG TCT TGA TGT TTG TCA GC | Wild type Reverse | A | |||
40447 | Fluorophore-1 | CTT TGT GTT TGA CGC AGG TTC | Quencher-1 | WT Probe | ||
40448 | Fluorophore-2 | CCC TGC ATA CTT TGT GAA GTC C | Quencher-2 | MUT Probe | ||
40449 | GTT TAT TTT TGC CTA CCA GTT TTT G | Mutant Reverse | A |
Component | Final Concentration |
---|---|
Kapa Probe Fast QPCR | 1.00 X |
ddH2O | |
40445 | 0.40 uM |
40446 | 0.40 uM |
40449 | 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 |