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Mutant =t/t
Heterozygote = c/t
Wild type =c/c
122 bp
GAGGCCCTAATGTCGGAACTGAAGGTCCTGAGCTACCTGGGCAATCACATGAATATTGTGAACCTGCTTGGCGCATGCAc/tGGTGGGAGgtgagtctcgagttgccatctctctggttatcaa
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
17087 | TTG ATA ACC AGA GAG ATG GCA AC | Reverse | A | |||
17089 | Fluorophore-1 | CGC ATG CAT GGT GGG AG | Quencher-1 | MUT Probe | ||
23173 | GAG GCC CTA ATG TCG GAA CT | Forward | A | |||
23174 | Fluorophore-2 | CGC ATG CAC GGT GGG AG | Quencher-2 | WT Probe |
Component | Final Concentration |
---|---|
Kapa Probe Fast QPCR | 1.00 X |
ddH2O | |
17087 | 0.40 uM |
23173 | 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 |