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>chr3:54367443+54367570 128bp GTGAAGGAACATTCTGGAAATC TCGGAGTAGTGCTGAGTGGT
Mutant= 116 bp
Wild Type = 128 bp
Fam=Mut
Hex=Wt
WT Sequence: gtgaaggaacattctggaaatcaacttctaagcatttgttcgattctgactCTttcagtgatgcctattgaccatgtttatggcacgctgggcattgtgggagccactaccactcagcactactccga
MUT Sequence: GTGAAGGAACATTCTGGAAATCAACTTCTAAGCATTTGTTCGATTCTGACTCaagctttctagggtacctctagctcgaggggcgcgcccccagctggttctttccgcctcagaag
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
12153 | CTT CTG AGG CGG AAA GAA CC | Mutant Reverse | A | |||
46218 | GTG AAG GAA CAT TCT GGA AAT C | Common | A | |||
46219 | TCG GAG TAG TGC TGA GTG GT | Wild type Reverse | A | |||
46220 | Fluorophore-1 | CAA GCT TTC TAG GGT ACC TCT AGC TC | Quencher-1 | MUT Probe | ||
46222 | Fluorophore-2 | CAG TGA TGC CTA TTG ACC ATG T | Quencher-2 | WT Probe |
Component | Final Concentration |
---|---|
ddH2O | |
Kapa 2G HS buffer | 1.30 X |
MgCl2 | 2.60 mM |
dNTP KAPA | 0.26 mM |
12153 | 0.50 uM |
46218 | 0.50 uM |
46219 | 0.50 uM |
Glycerol | 6.50 % |
Dye | 1.00 X |
Kapa 2G HS taq polym | 0.03 U/ul |
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 |