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Taqman qPCR protocols are run on a real time PCR instrument. Use an appropriate instrument specific Fluorophore/Quencher combination.
Mutant= 105 bp
Wild Type = 105 bp
>chr6:106781062-106781166 105bp GCCTAGTACAAAGGCTAAAAATAGG TCCAATATGGCTTGCACAGA
Sequence: aacagcaacatatagacacgtgcagtaataaattatccagttataacagggtggctttgtaaatttgaggtgtcagagaaatctcacgagaagcctagtacaaaggctaaaaataggttttaacttatatctttccttcattttatagGTATGCATGGACC(A/G)t(t/g)GCCCAGTGCAAGATCTGTGCAAGCCATATTGGATGGAAATTTACAGCCACAAAAAAAGACATGTCACCTCAAAAATTTTGGGGCTTAACTCGCTCTGCTCTGTTACCCACAATTCCAGAGACTGAAGATGAAATAAGTCCAGACAAAGTAATACTTTGTTTATAAGTGCACCTGTAGGAGT
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
49562 | GCC TAG TAC AAA GGC TAA AAA TAG G | Forward | A | |||
49563 | TCC AAT ATG GCT TGC ACA GA | Reverse | A | |||
49564 | Fluorophore-1 | CAT GGA CCA TTG CCC AGT GC | Quencher-1 | WT Probe | ||
49565 | Fluorophore-2 | CAT GGA CCG TGG CCC AGT G | Quencher-2 | MUT Probe |
Component | Final Concentration |
---|---|
Kapa Probe Fast QPCR | 1.00 X |
ddH2O | |
49562 | 0.40 uM |
49563 | 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 |