Taqman qPCR protocols are run on a real time PCR instrument. Use an appropriate instrument specific Fluorophore/Quencher combination.
Mutant= 104 bp
Wild Type = 101 bp
>chr5:117203488+117203588 101bp ACACACATACGCATGCACAG TCTAGTTGTCTGGGAATAGATAGCA
Sequence(deletion in upercase) ttagtgcctcggttggttggttgtgtgcttggttgattggttagtcagttggtcgattgggtacttgggtaggcatatggctggttggttggttggttggttggttggttggttggttggggtaggttggattgtgattggttgcttggttgattgattggctgtttggttggttttgtagctatttggttggtttggtgtggttAACTTGAGGAACTTCTCATTAACTTTTATTATTCCGATCTTCCACTCTGTTCCAGTTGGTGATGGAGTACTGCCTAGGTTCAGCCTCTGACTTGTTAGAAGGTAAGCATGACATCATAGCCTCTTCTGTTTTCCTTCTGTCACGTTCTCCATGGGGCTGGAGAGACAGCTCAGCGGCAAGAACACTCTTCTCTCTCAAGGACCCAGTTGGGCTCCCAGGCAAATGCCTGTAACTCCAGCTCTGGGGACCTGGCATCCTCTTCTGGCCTCCACAGGCACCTGCACCCAATGTGCACACACATACGCATGCACAGGCAGGCCACATgtacaggcacggacactaaaataaaatcttaaaaacaaaaaagaatgctatctattcccagacaactagacatagtgcagtttgagcctctggttgaaaggcagtttaagggtcactgggtatactgtatagacataatctctacactatagctgggtgatctcagctgcatggctgtcccgggcgggcactagccaaaaaccatcaaagccatgtataaatgtcactgcctgtgcctttccagtatcacagtgggcacggatgtgagaggggaggtgctgtcaactcactagtatctacccatgtgcagagtcaacgacggacctgagctctcccaccactgctgctgtgtgtgcccagcagctagctgtgtttaaaaattctattta
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
40576 | GGT TTT GTA GCT ATT TGG TTG GT | Mutant Forward | A | |||
40577 | TCT AGT TGT CTG GGA ATA GAT AGC A | Common | A | |||
40578 | ACA CAC ATA CGC ATG CAC AG | Wild type Forward | A | |||
40579 | Fluorophore-1 | CAG GCC ACA TGT ACA GGC ACG | Quencher-1 | WT Probe | ||
40580 | Fluorophore-2 | TGG TGT GGT TGT ACA GGC ACG G | Quencher-2 | MUT Probe |
Component | Final Concentration |
---|---|
Kapa Probe Fast QPCR | 1.00 X |
ddH2O | |
40576 | 0.40 uM |
40577 | 0.40 uM |
40578 | 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 |