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Mutant = GCCGCC/GCCGCC
Heterozygote = AAGAAG/GCCGCC
Wild type = AAGAAG/AAGAAG
CGCCTCCTGCGCCATCCAAGTCCCCGGAGGAGATGCGGAAACTCTTCAT
CCAGAGAGCC(AGGAGG/GGCGGC)ATCGACAAGAT(a/c)TCTCGCATCGG
TTTCCCCATGGCCTTCCTCATCTTCAACATGTTCTACTGGATCATCTA
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
18234 | GTG CCA ACA ACA ACA ACA CC | Forward | A | |||
18235 | ATC CTC TCT CCG GAC GAT CT | Reverse | A | |||
18236 | Fluorophore-1 | AGA GAG CCA AGA AGA TCG ACA A | Quencher-1 | WT Probe | ||
18237 | Fluorophore-2 | AGC CGC CGC CAT CG | Quencher-2 | MUT Probe |
Component | Final Concentration |
---|---|
Kapa Probe Fast QPCR | 1.00 X |
ddH2O | |
18234 | 0.40 uM |
18235 | 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 |