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A standard PCR protocol has been published and can be viewed online at doi: 10.1093/biomethods/bpaa007. Please note, the assay in this publication has not been validated by The Jackson Laboratory.
Mutant = Deletion of highlighted G
Wild Type = Presence of highlighted G
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
15131 | Fluorophore-1 | ATC TGA TGA CAA GAC TGA GTG TTC G | Quencher-1 | WT Probe | ||
15132 | Fluorophore-2 | ATC TGA TGA CAA ACT GAG TGT TCG | Quencher-2 | MUT Probe | ||
54913 | GCT GGG TGT CAA CAA TCA CA | Forward | A | |||
54914 | AGT CAG AGG GCC CAT ACA GG | Reverse | A |
Component | Final Concentration |
---|---|
Kapa Probe Fast QPCR | 1.00 X |
ddH2O | |
54913 | 0.40 uM |
54914 | 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 |