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Mutant = 297 bp
Heterozygote = 198 bp and 297 bp
Wild type = 198 bp
Ignore floxed band at ~250 bp for scoring purposes.
Primer | 5' Label | Sequence 5' → 3' | 3' Label | Primer Type | Reaction | Note |
---|---|---|---|---|---|---|
26219 | GTA GAC GTG GGG ATC AGT CC | Common | A | |||
26220 | CGC TCT GCT TTT TGA TGT TC | Wild type Reverse | A | |||
26221 | ACT CAC ATC ATG GTG GAA AGC | Mutant Reverse | A |
Component | Final Concentration |
---|---|
ddH2O | |
Kapa 2G HS buffer | 1.30 X |
MgCl2 | 2.60 mM |
dNTP KAPA | 0.26 mM |
26219 | 0.50 uM |
26220 | 0.50 uM |
26221 | 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 | 94.0 | -- | |
2 | 94.0 | -- | |
3 | 65.0 | -- | -0.5 C per cycle decrease |
4 | 68.0 | -- | |
5 | -- | repeat steps 2-4 for 10 cycles (Touchdown) | |
6 | 94.0 | -- | |
7 | 60.0 | -- | |
8 | 72.0 | -- | |
9 | -- | repeat steps 6-8 for 28 cycles | |
10 | 72.0 | -- | |
11 | 10.0 | -- | hold |
Stock Number | Strain Name |
---|---|
028040 | B6(Cg)-Fxnem2.1Lutzy/J |
029100 | B6.Cg-Fxnem2.1Lutzy Tg(Ckmm-cre)5Khn/J |
029720 | B6.Cg-Fxnem2Lutzy Fxnem2.1Lutzy Tg(Ckmm-cre)5Khn/J |
030218 | B6.Cg-Pvalbtm1(cre)Arbr Fxnem2.1Lutzy/J |
029721 | B6.Cg-Pvalbtm1(cre)Arbr Fxnem2Lutzy Fxnem2.1Lutzy/J |
031007 | STOCK Fxnem2.1Lutzy Tg(FXN)Y47Pook/J |
030395 | STOCK Fxnem2.1Lutzy Tg(FXN)YG8Pook/800J |
030930 | STOCK Fxnem2.1Lutzy Tg(FXN)YG8Pook/J |
8 strains use this protocol |