Taqman qPCR protocols are run on a real time PCR instrument. Use an appropriate instrument specific Fluorophore/Quencher combination.
Mutant= 114 bp
Wild Type = 108 bp
>chr8:110841833-110841940 108bp CAATTGAGAAGCAGAAATTGGT CGTGATACACCAGAGTGTTCG
Sequence (deletion in uppercase)
tttcttagtttatctcatcttcaatagatgattaatgatcttatttaaaagaataaatatttaggtttctgtcttttcatgaaatctgctactttatctcctttaacaacaggtgcaattgagaagcagaaattggtatatatcttgaacagagacgctgcagccCGACTTACCATTTCCTCTCCCCTGGAGGCCCACAAAGCGAACACTCTGGTGTATCACGTGGTTGGCGTAGACGTGGGATTTGAGAATCCTATGTTTGCTTGTCTGGAAATGGATTATGAGGTACCAAGGTGTCTCCCTCCCTCCTCCTCCTCCTTGGCTGTTTTCGTCATGTCCGATTTTTTCCTAAAACAAGTTTAATTGTGTCTGTAGATCTTAAAGTCATTCTTGCTAATGGATCTTTTTCAAGATATGAATGCTTTACTGAGAGTCTTTGCTGCACTTGACTGTGAAGTATCATCTGACTTGAATGACTGGACTGGGTCTGACCTGAGATATTTATTGAGAGGAGCTGCATGAGAAAAGCAGAGTTGTGTGTGGGGTTGGGGTGCTGGTTTGTTTAAGGGGAGCAGTAAAGGCTGAGCACACCGCTCCTGGCTTGGACGGAAGGCAGTGTTCTCTAGCAGTGCCCTGCCTGGGTCCAGAGCAGAGATTTGATCCtcgtggttctttctacctttgtagcttaaactttgtcaaactatttgcaatctgtgaacctttgcttcactgtatgtaaagtcctgatatcctttccaagtctaaaacttgatgattcgtttctgtggcactgtct
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
40581 | CAA TTG AGA AGC AGA AAT TGG T | Common | A | |||
40582 | CGT GAT ACA CCA GAG TGT TCG | Wild type Reverse | A | |||
40583 | CAA AGG TTC ACA GAT TGC AAA TAG | Mutant Reverse | A | |||
40584 | Fluorophore-1 | CGC TGC AGC CCG ACT TAC CAT | Quencher-1 | WT Probe | ||
40585 | Fluorophore-2 | CGC TGC AGC CTC GTG GTT C | Quencher-2 | MUT Probe |
Component | Final Concentration |
---|---|
Kapa Probe Fast QPCR | 1.00 X |
ddH2O | |
40581 | 0.40 uM |
40582 | 0.40 uM |
40583 | 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 |