Taqman qPCR protocols are run on a real time PCR instrument. Use an appropriate instrument specific Fluorophore/Quencher combination.
>chr8:105086275+105086382 108bp TTTGAACAAAGGGTGTTGTCTC TAGCAGGACAGCCAGTCAGA
Mutant= 109 bp
Wild Type = 108 bp
AACCAAACATGAATGTTGAATTTGAACAAAGGGTGTTGTCTCTAACTGGGGCTAGAgtcctagcagggaatgaggacaaagagccatcttcaccatctaagagccagctctgactggctgtcctgctacatctacccacaacacatactaggaaaacaggtgattcacgggcaagcaaggactgggaagaggagaagtgcccagtgccctggtgactgactcaaagcaaatgcacagagtgtaagaaaaaaaatatgtcttggacgctatcttaagccaatcagtgccactccctgtccctagcactggagacaagcacatgccaggcaacaggggattcctggatgtggtggctgctcttcgctgggtccaggggaacatagccccctttgggggtgatcccaactgtgtcactatcttcggtaactctgctggaggcatgattgtctcatccctggtgagtgaccacagagaactagcagtcgaggttatatctgggaaggcttctatggcctcagacctccatgctcctctgcctcctaacatctgcccctctgttgttcccagttcctgtctccaatatctgctgggctcttccacagagccatatcccagagtgggattgtcacgaccattatgatggaagacatgaaaccatggcctgaagctcaggtctgcctttccttcactctcttaccctaagtgtgcctccacagcccaccctgtgctaacctttgggtcaggtgtggagaggaaggagtaaccactagggagtcactgataggctcagagacaattggctagtgcagtccaggaggtaggcagtcagacaaggctttctagaggaagccaccctggaagaggttatctctggttcttctctggggttatttttgtagatcccccccattccccaacgGATTCCTATATTTCCAGCTACTCTGTTACCTAAGAGTTCGGTCCCCTTTTGAGGAGTAGCCAGAAGTCACCAAGCAATCCCTAATAGGAACTGTGAGTCAAAAGCTTCATGCATAATAAGACAGGGATGGGGCACAGAAGCAGTAAAGCCATCAGGTAGGGAACAAGCAGGAAGTGAATGAACTAAAGGAGTGGAAGGTAGTGCAGGGAGCTGACAGACCATGCCTGGGAGGGAGTTTGGCGGGATGCAAAGGGCTAGAAGAAGCAG
An 863 bp deletion beginning at Chromosome 8 position 105,086,311 bp and ending after 105,087,173 bp (GRCm38/mm10).
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
45670 | TTG AAC AAA GGG TGT TGT CTC | Common | A | |||
45671 | TAG CAG GAC AGC CAG TCA GA | Wild type Reverse | A | |||
45672 | TTG GTG ACT TCT GGC TAC TCC | Mutant Reverse | A | |||
45673 | Fluorophore-1 | TGA GGA CAA AGA GCC ATC TTC | Quencher-1 | WT Probe | ||
45674 | Fluorophore-2 | CTG GGG CTA GAG ATT CCT ATA TTT C | Quencher-2 | MUT Probe |
Component | Final Concentration |
---|---|
Kapa Probe Fast QPCR | 1.00 X |
ddH2O | |
45670 | 0.40 uM |
45671 | 0.40 uM |
45672 | 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 |